ELL, CTRICrisiill,,,Vj-:STIGATIOI-4 -Fin.,ii Report coiitract 8 llarcil 1973 ILLUSTRATIONS Page 1 Mautluier cells 4 2 Electroreceptors 6 3 Cranial iiel-ves of Eleci@-.-i)pliorLis 8 4 Microelcctrode amplifier 13 13 5 Microelectrode amplifier. schematic dia,-ram 6 Loss in -dB with increasing f2requency 14 7 Loss in -dB with increasing repetition rate 15 -.8 Device for sharpening metal electrodes 17 9 Device for applying silver chloride 17 '10 2 Potentiomet,ic device 19 11 Device for me,-tsuring niicroelectrode resistance 19 Amplification factor, square wave 20 11A 11B Amplification factor, sinusoidal wave 2 21 iic Recordinos at different square wave 22 11D Recordin"s at different square wave 23 12 Adjustable Lticite Tray 27 13 Instruliientatioii for Anaesthesia 2 27 14 Solution 1:10000 in water on Stcrnarchus -Lll)ifrons 29 15 Sterii,,irclius .ill)ifrons i'@7 normal electrical activity 31 16 Sterii-,trclitis all)if-rons #7 in anesthetic 2 tricaine inctliaiie 31 sulfoiiate, I miiiute 17 Stei-ii.,trclitis ,ill)ifroiis llr'7 in,,tncstlictic tricaiiie mcthmic 31 sulfonatc, 2 mbiutes 18 Stern,,irelius .ill)ifroiis 1!7 in.,ti-icsthetic tricflille rLICtIl"lllc 31 sulfoiiate, 4 minutes 19. StCrll-,Lre-Iiiis -i.ll)ifroiis lr,7 in anesthetic tricaine metliaiie 32 sulfonate, 5 iiiiiiutcs 9 ?O. reciil)cr.-itin,-, 5 ii-iintites 32 I>a4rre 21 Stern,,irclitis all)ifroiis recuper,,Lting, 8 minutes 32 32 2 22 Set-up for microciectrode 23 Stern-irclitis -tll)ifrons No. 1 just before amestliesia 33 24 Stern,,trclitis ,Lll.)ifrons No. 1 maesthelizcd witli ti-dopental sodium 33 (2 mili. 2 Stern, -lietized wi i thiopental sodium 34 25 irelitis qll)ifrons No. 1 anaest II (13. 5 min.) 26 Sternarchtis albifro-ns No. I anaestlietized witli thiopental sodium 34 2 (37.5 min.) 27 Sternarclitis albifrons No. 2 in aquarium water 36 28 Sternarclius -dbif-rons No. 2 in MS-222, 10 minutes 36 29 Set-up for recording 2 38 30 Close-up of the set-up for recording; 38 31 Sternarchus ,Lll-rfj-oiis after d-tabocurarine injection 40 32 Ampullary, tonic electroreceptors 40 2 33 Set-up for recording 41 3.1 Anaestlictized, curarized Stei-iinrchtis albifrons 41 35 Microelectrode amplifier 43 3G Preaniplifier 2 44 37 S3,iiclironous tonic electroreceptor recording 44 3S Nonsyilclironous pliasic electi-oreceptor recording 44 39 Gvmii-.irclius nilotictis Cuv. #2 electric activity 44 .10 GNiiiiiirrlius niloticus Cuv. #3 electric activity 46 -11 Cviiiiiirchus niloticus Cuv. Tlr2 electric activity 46 -61-1 Cvi2iiii.,trclius niloticus Cuv. 1'1"2 electric activity 46 Civmitircliu-, niloticus CtiN,. #3 electric activity 4G Cl-vilill'ircliiis iiilotic,lis Cuv. i';-3 electric 'Lctivit-.y 47 k;vii ii-.irelitis nilotictis Cuv. 113 electric activity 47 C-viiiii-irclitis iiilotictis Ctiv. (baby) - tr-"nsvcrsil cut 48 iv F inii r e 47 G_Ini-inarchus niloticiis Cuv. air bladder and spinal cord 49 49 48 gvmnarchus niloticus Cuv. - spinal cord 49 Gyn-iii,.xrchtis niloticus Ctiv. - brain 2 49 50 Delafield - Harris hematox-ylin staining method 50 51 Malaptei-urus electricus -t2 52 Malal)tcrus clectricus on the scale 53 53 Device for measurin,-, the2 voltage 53 54 Discharges of the electric organ of Mal rurus electricus 54 55 Tonic, ainpullary electroreceptor 56 56 Phasic, tuberous electroreceptor 57 57 Tonic electroreceptor 2 59 '60 Phasic electroreceptor 59 Ampullary tonic electroreceptor 61 60 Stimulus recording 61 61 Tuberous pliasic electroreceptor 2 61 62 Stimulus recording 61 63 Electroreceptors of G@vin-narclius niloticus 62 64 Under-,vater pattern recognition system 63 65 Recordin-, from an anaestlietized, 2curarl--ed 66 Sternarclius -.dbifroiis specimen 66 'Alicroelectrode recordincr 66 67 Gymiiarclitis niloticiis - photo 70 68 G)n-iiii,,trclius ni.2lotictis - placeincnt of electric 71 transiiu'tilii- micl receiviii- org,,tns 69 Fish L,,tl)oratol-y A 72 70 Fisli L.-il)or,.itol-y B 72 Tnl)lc pittcrii rcco,,-nitioii system' 55 v SUMAIARY The electric or,,,,,tiis of Stei-ii,-trclitis ill)ifro)is, a South American fresh water wealt electric fish, li,,t-%,c been studied with emphasis on clectroreceptors. The morpholo-- 0 ic-al and i)hysiolo,,-ical characteristics of clectro@-eceptors, ampullai-Y and tuberous, a.. e discussed. Special instrumentation required for the role of these electroreceptors in pattern reco-i-dtion has been developed. We have recorded with microelectrodes the autonomous au2torhythmic electrical activ- ity of the tonic as3,nclironous aiiipullary electi-oreceptors of the South American weak- fresh water electric fish Sternarchus all)ifrons. NVe have also recorded the elec t-rical activit@,, from the phasic tuberous electroreceptors and of the synchronous an-ipullary electroreceptors of the same electric fish, Stern-,irchus albif-roiis. Preliminary meas- urements have been made . The electric dischar,-e of i'@Ialai-)terurus electricus, an African fresh water stroii(r electric fish, has been measured in and out of water. The autorliythmic activit@, of the ampullal7 electi-oreceptors has been demonstrated. We obtained some specimens of the African weal.: fresh water electric fish G@,mi-iarchus niloticiis. They are supposed to be the most sensitive of aH the weak electric fishes k-nown. Together with two specimens about one foot Ion-, we received a number of baby GNmn,-irclius nilotictis ,tbout two inches -loiicr. TIie baby electric fish were in- fected wi2th a Saprolc,,,qii-,i fimgus and could not be savcd, but we fL-,@ed a number of them in buffered formaldehyde and one of them has been cut and mounted in paraffin for histolo-ic,-LI studies or the electric or-ans. Preliiiiin,-Li-y meistii-cments have been made on the coiiimuiiic.,Ltioi2i cipal)ility of .idtilt Gi,iiiii-,irelius iiiloticus. A stud3, of the iiiestliotiziii,r effect of tricaine-iiietb-.tiicstilfoiiite (AIS9-22 FINTQUEL) LI on Sterii.-trelitis -tll)ifroi-is liis I)cLii Lii1ldcrt,-J@cil by plottiii,-, tinic for the anesthesia and I recover3, for difiereiit spcciiiic)is. vi A study of an -,inaestlictic which does not affect the electric fish's electric org'.In pulse repetition rate is presented. Also, the effect of D-tiibocurariiie and the cotintcr-effect of neosti-mine has been LI assessed for Sterii-irclius alb2ifrons. r-inally, some impronements in the micro- electrode recording instrumentation have been made. The electric orcraiis of Sternarchtis -tlbifrons a South American wealc fresh water -udied with emphasis on clectroreceptors. Recordings electric fish, have been st have been made from the asynchronous tonic as well as the synchr2onous tonic and the asynchronous phasic electroreceptors. The electroreceptors are part of the complex lateralis line system of the electric fishes. The other lateralls line system sensory receptors, lil,.e mechanical receptors and displacement receptors, have been discussed as part of a general hybrid pattern recog-ni't-ion system of the fish. A passive hybrid tm2denvater pattern recognition simulation syste'm has been advanced. A simul,-ttine, model concept could be established for underwater patt,-rn recognition throu,- .,Ii electric sensory receivers and electric fields. More histological worl@: is needed to establish the r,-,Iatioiisliip between different electi-oreceptors and their innervation. This is also needed for a realistic simulation system of the underwater pattern recognition ability of the electric fishes. I. INTRODUCTION En one of our previous reports, we described the morphology of the electric organ Df Sternarchus all)ifroiis, a we,-d@ fresh water electric fish from South America. We mentioned that the electric transmitting organ of Stei-narchus is derived from nervous tissue and no2t from modified muscle tissue lilce the majority of other elec- tric fishes. This is mal@iiig Stcl-ii,-irchus different from other electric fishes: it has a very 1-d-h sic ID ,q-lal r,-tte and the signal is phase and amplitude modulated. The form, rate, and amplitude of the signals emitted by the el2ectric fishes are as diverse as the forms and sizes of these fishes. Some of the wealc signals are used to locate- objects or animals in their environment, or for navigation, species recog- 2 nition, and communication. The strona electric discharges serve for offense or 3 defense. IVatanabe and 2Tal@eda in,%resti(rated the effect of a-c current with a fre- quciicy close to tl-ic electrical signal en-Atted by Ei,-enmanilia. When the applied pulses came within + 3 to 4 pps of the one eii-iitt,-,d by Ei,-enmal-u-Aa, the fish would chancre its rate by 4 to 5 pl)s in a direction 2xvliich increased tfie pps separation. In- creasing the frequency of the applied a-c current in 1 pps increments caused the fish to shift its frequency correspondingly until it reached about 6 to 7 pps over its normal rate, when it would rei,ert to its orizinal rate. We obtained similar results in c.%Wriments with Steni-,irclitis ill)ifrons, but the applied a-c signal was within 0. 5 cycle of the signal of the fish, clemonsti--,ttin- how specific the ,Lpplied signal must be to elicit a chan,,c. in the Stc).-iiirclitis si-lial r,,ite: + 0.5 cycle is the range of se- 2 lectin-ity of the fislies' electrical trmismittinfr-receii,in- system. The fact that many 0 0 fresh water -tlld set N\,.-itcr electric fishes h,-L-%,c never licen stu(UcLl may present some (lifficillues 5in ol)t.,Linin,- speciil ]@iii(Ls of elccttic fishes, and their care ni,.iy not be C3, "ii easy tlisl@. There irc, liowck-er, ciiou-Ii species to ciial:)Ic iil-.Iii3, c.%-pcriiiient@s. Very little is luion%,n 'LI)OLIt the electrical activity of marine electric fishes except 12Mcdo -Lnd some rays. N,,ireine 13-irziliensis is tlic- only Izio-,%,ii marine electric fish having two different electric organs. Bemiett 4 studied the mode of operation of2 the electric organs of R,,tla c,-I,,LnLtria, a marine ciectric fish, and of the fresh water fishes 11@,-popomus and Stcrnol)y-,iis and compared them with N,.trcine, l@lormlrrus, Steato-,cii3,s, Gymi-iorl-i.-L M,-il,-tpi2crurtis, G3rmnotus car,,il) ,and Electro- The main objects of this study were the form, innervation, and physiology of the electi,oplates formin- the electric transmittin- organs. The clecti oplates of. Hypopomus, Gyliinottis carapo, M-,ilal:)tcrurus, and some Mormy-l-idae have the same 2 surface area and produce spil@:es durin- discharge. The electroplates of Stenlopylrlus and possibly Eigenm-.tnnia have tino peculiar characteristics: there is a steady po- tential on which pulses are superimposed, and the resistance of the electroplates is similar at the peak or between the spikes. The electroplates are of the type with a 2 slow depolarization. In IiL-popomtis, l@lal,,tpteriii-tis, ,tnd most or the Morm@Tidae, the inner,,,,ation of the electric organs is through stalks. The stall@.s may serve to amplit, depol.-Lrization until it would be able to invade the body of the electioplates. The disc2harge rate -,tnd duty cycle have been compared for electric fishes like Stcrnol)y,ni (rate = 50/sce) and Eigcnm-,iiinia (r-,ite = 286/sec) with the Stemarcllidae (mo.%. 1500/sec). Compared with mamm.-tli,-ui central nervous systems, peak- fre- ciucncies of the electric orc2riiis .ire not greatly different. The R-,iishaw coR can cliscliarcre impulses ,it a rate of 1400/sec5 and neurons in the sensory path some- 3 6 Linlr-,s produce btirsts at a rate of ibout 1000/sec. C%,,,tminitioii of (liscli-,ir-c'l).-tttcrn and or,-,mi function studies, it was noted that C. 0 Crtui(ifest7deriiicd tivo grotips of electric fislics: those that emit sigm-.ils at a constant 8 2 rate, and those that emit pulses with a variable rate. r- or example, Gyr=,Lrcbus, Sternopygus, and Ei-cnmamii.,t,,tre in the constant rate group; Electrophorus, Gr,ithoneiiius potersii, Steatorron3rs, and I-Iipopomus belong to the variable rate 4 group. BemetL 2did not malze any connection between the electric fishes' electrical systems, their environment, and their belianior. No one investigated their evolu- tion, very little is known about their matin- or birthplace, and no one has reported the broodiig of electric fishes confined in water tai@@. Sternarchtis albifrons also has two kinds of e2lectroreceptors: tonic and phasic, and they are autorliythmic. These electroteceptors are sensitive to movement and di- rection. The phasic electroreceptors seem to be related to informatioa regarding movement of objects near or around the fish.. Acceptin- the principle of pacemaker activity in the brain, it seems that there are only a reduced number of command nuclei acting on the 2electric transmitting organ. It is also reasonable to assume that electrically mediated positive feedback must be present; chemically mediated transmission would be too slow for the repetition rate of transmission which can attain under certain circumstances over 1, 300. Alautlmer cells of lower vertebrates (Figure 1) can be considered single cells 2 8,9 command system for the, axial musculature on either side of the body In the hacketfish each Mautlmer fiber activates the muscles depressin-S both pectoral fins t@- and these cells thus constitute a bilateral command system for the depressor 2 10,11 muscles. For e%-plaining the pacemakers action of the command nuclei in the brain of the mormyrid electric fishes a mutual excitation with positive feedba-ci@: has been proposed. This theory would not work for the Mautluier cells. There is a crossed inhibition between the Mautlner cells in the brain of the goldfish and it could equally 9 be effective in an electric or-an system. There is a requirement of 1-d-h speed 'If transniissl'o'n in s@nchronized systems like many of the electric tr-.insmittin- 0 organs. This has been useful in predictin- sites where tr."siiiission has bccn JL -J 0 CD SL 0 CD -- - ---- - ---- electrically mediatecl. Positive feedback may be a "sinc cltia non" requirement for the pacenial@er nuclei of electric transmittino- organs. There is proof of positive fced- 12,13 back in the mutual inhibition system of arthropod compo2und eyes Th) neural systems controlliiig electric organs have provided a large number of ex,.mples of electrically mediated transmission, which meets the functional require- mlnt for rapid communication between cells. This mode of transmission also pro,.,es t to be able to mediate many functions often considered as restricted to chemically mediated t2ransmission. The correlation between morphologically close apposition and electrotonic couplina- was considerably strengthened by the work on electro- motor systems. This correlation helps to validate morphological identification of electrical transmission in other systems where electrophysiological analysis is not so simple, It is not known whether there is any relevance 2to hicrher systems of the organizational principles deduced from electric organ systems. The next level of analysis of the electric organ systems may be no easier than the study of less specialized systems that are of more general interest. Some kmowledge is being obtained of afferent pathways from electroreceptors in weakly electric fish which have important inputs to 2the electric organ control system. Both operant and respondent conditionin- of the control system can b@ obtained and conditioned response latency can be very short. It is not unreasonable that the complete neural pathway of the conditioned response could be obt,-ined in these cases. The central connections are still minimally explored; one knows what -oes in and one can go from 2the electric or,-an SCI,eral synapses mitidromically. The rewards for fillin- in the gap would be great, and prospects for at least some pro-ress are bri-ht. In.sternarchtis all)ifrons clectroreceptors are distributed o-%,cr the entire body. The 'S'rccol)tors are vl-ry numer8ous as conipare i -asic tul)@--rou d with the tonic amptillai-y rec!CPtors (rl-ure 2). The density of receptors is greatest in the head re-ion ind 5 pullary Receptor Tonic, Am v iI v for 2 f Phasic Tuberous Receptor 'V v ... ou ter' 2 fac r ne -face rnal, and 2 receptor externc3l, int cell skin resistances gil i,c 2 Ail:itoiiiic-al dia-i-.-tms -inci cqtii\-.ileiit circiiits of cicctroreceptors in 2 frosti watcr clecti-i(! fisli: (a) toiiic, ai)il)tillary receptor an(I (1)) pli-.isie, Itil3ci-otis rocci)tor. Diitrr-iiiis are slikl\vtl witli ilic cxtei-nil iii2(,diulii to the top. 'I'lic si@in .ui(i wall of i-c(,cl)tor ea,%-it ics :ii-(,. Rlio%%-ii iii cross 'I'lic tipeiiiii- to Llic c.,cterior of 1)liisic c. -ity s( etioi as Iiiies. '1'% is sliowii is o(-citi(lecl ]-)y -i poi-otis iiia.9s. 'I'lic iici.,%-c fil)(!rs iiiiie-i-5\,:Ltilig" receptoi- ,tre iii(li(-.ited. falls gradually to,@%,ard the posterior end. There are minor morpholo-ical subdivisions thin the phasic and the tonic receptors, but no physiological correlations have been %Vi yet obtained in r, icrure 2 the equivalent circuits of tonic, ampullary and phasic, tuberous organs is shown. Cross sections tlirou2,- ,h the receptors are shown with the external medium to the top. The skin and walls of the receptor cavities are represented by lines, innervation of the receptor cells is indicated. The electroreceptors over the entire body are innervated by the anterior lateral line nerves, a large branch of which runs posteriorly to join the posterior lateral2 line 17 nerve just behind the head (Figure 3). The posterior lateral line contains only 18 19 chanoreceptive fibers which come from free neuromasts and canal or-3ns me 'nie receptor cells of tonic receptors appear to behave very nearly like linear 2 elements; that is, their membranes have fixed internal potentials, resistances, and capacitances. They are, in a sense, electrically inexcitable, and they differ markedly in this respect from phasic receptor cells. There is evidence for chemically mediated 20 transmissions at tonic receptors of -viiinotid The morpholo- cal characteristi2cs 0 se f chemically mediated transmission. A strong f the snyap are those typical o brief anodal stimulus produces an evoked response lon- outlastin- the stimulus. Then there is a synaptic delay between the initiation of the impulse and the nerve impulse (between 0. 5 and 1. 5 msec). Mormvrid tonic receptors are similar to thos2e of gymnotids and Glmn-,trclius nilotictis tonic receptors are morpholo-ically similar, 21 but they were not pliysiolo-ically studied The relationship between the different electroreceptors of Sternarchtis -,ilbifrons in pattern recognition has not as yet been studie2d. The present study developed special instrumenll-ition required for in-%,esti-,,ttin- the roles of these elcctroreceptors in P'Ittern reconiiition and obtained preli 2surcments of electrical discharges f roni Is in ind out of water. 27 ;,L F i gii rc 3. liorizoiit:tl 1)rojectioii of the craiii-il liern,cs of lIccti-ol)liortis. Notc tlic! siiiill siz(,! of the br.-iiii. Sonic ncr\,cs -,ire in(licated I)y tfic coi-rcsi-)oii(Iiii,,- nunil)crs. L.A. iior,,-us latei--.ilis -iiiterior; -us Iitoi,-,ifis postc.-i-ioi-; Oc-SI), ocvil)it-o-sl)iiialis iierve. 2 LI" iiern In our e.%I)erimcnts, the perception of ol)'ccts by weal@z electric fislic-.; b.N'w-'LY Of the I i (Icmonstrated. discharge of their electric organs (transmitting an2d receinring) has bcci 22 "u tuthors have shown that wealc electric fislies'can be trained tu distino ish other between conductilltr and nonconducting objects placed in the water. For this Idnd of reception, t-%io possible modes of action at the level of spccific re- 22 ceptors could be proposed: the "pulse-frequency-modulation!l and the "pulse- 23 phase-modulatiol@' According to the fi2rst hypothesis, sensory information should be conveyed by the frequency of the sensory impulses dependent on the pulse Of the electric organ discharge Nvl-iereas, according to the second hypothesis, the time re- 'OlY i lation (the phase2) between the electric discharge and the following scii.@j mpulse ciits with would play an important role for the sensory codin,-,. From our expcrim tr Ste2rn-,Lrclitis ,ilbifro)-is a South American wealc fresh water elee ic h, we con- M eluded that apparently neither of the above proposed mecha-,iisnis are operatint2 2 coded by this fish. The intensity of current flowin- at the level of the receptor i-S the number of impulses elicited by each electric orgaii clischar-c. S-tl-.-rn--rchlis albifr2ons can dischar-e at rates higher than 1000 per second and the 5@nnsOrY impulses rs cali follow their discbar-e rate. of the recepto 24 25 In some mol-ni3,rids and g3,iiinoti&- however, modulation of the rr;lation electric- 2organ pulse-sensory ii-npulse may be used for electrosensory codiiicr. Li mormyrids, 'the cliaii,-- also, -ling _,c in intensity of the electric field may be coded by cb the atency 2 to iL ds In morm Ti bct,,veen the electric or--.tn pulse and the sciisol-@r recep rs' imp ilse. it niay attain values til) to 9 -n-isec, whereas iii gyiiu-iotids it has been f,,-.in-d not to exceed 1-2 msec. It lias been previously ii@clitioiicd that in Stern,,Lrclius albifroiis the el(,,;-,romeeptors distril)utc(l over the entire bo(ly of the fisli md tli,-it the anil)ullal7- 2 c -ire rd receptors niore niiiiicrotis tli,,Ln the till)crous ph-.tsic receptors. we rccordc:,.' -.-,Qn the ,Luto- rli@,tilll-de clectricil ,Lctiviti, of the iionsyiicliroiious toiiic, ,tniptillii-3, ---;"rorceeptors Of Stel-ii-irclitis ill)ifi,oii.-3. Tllc iiiipulses were irro,,,Yiil.-.r -irottii(I t rtite of bet%vccn 100 ,tnd 300, with an amplitude of around 2.5 mV. The impulse duration was around 200 microseconds. There ,ire other types of tonic receptors which are -onous. synchronous. The phasic units are noiisyiichl The non2synclironous tonic receptors seem to react independently from the trans- n.,iWn,-, electric or-an. They react to any objects brou,- t near the fish at a certain h range. The recording shown in our final report is made from such type of receptors. 2 As previously mentioned, some fresh water weak electric fishes have the ability of perceivin,- objects, their movemenf and direction, and also to determine some char- acteristics of tlicse objects (such as conductivity). For tlds underwater pattern recognition, they utilize both their electric transmittincr organs and their electro- receivers. Obviously some other sensory perception receivers of the lateraus line may be involved such as: free neuromasts and the ctipula type of lateral line re- ceivers; the auditi-%,e system may play a role also in this pattern recognition. Some of the electric fishes are blind2 or have vestigial eyes; others liaie good vision and it is certainly used in the reco,,m, iition process. It seems that a hybrid system made of a diversity of sensory receptors is used by the electric fishes to locate and identify objects and fishes of the same or different species and also prey or predators. They use little elec2trical energy for this and the distance involved is coiisideral.)Ic - it may attain, under certain circumstances and for one specific sensitive species (G@-niiiarcl]i@, over a mile.' The sixoii.- electric fishes (Elcctrolllior-,is, Tol-I)cclo, and Astros2coptis) c.,tn incalmcitaw md ldll their prey with their discharge of the powerful 3lectric org,,ul used only for such I)tirl)oses. If they have a detectili,- and loc,'Itin- system, thc3, use ,L different low power clc2ct3.,ic ta-.-uisniittiii- than the main powerful electric C. The meaning of this is ili.,tt there is no Iii-h power I'Oqtiireniont for the pittcrn reco.- ,iiitioll system of electric fishes. 2The electric 10 sensory rcccl)tors ,ire very sciisiti-%,c, have a lij,-h discriminatin- captcit-.v, and use an electronic proccssin- system I)ascd on a niultil)lc degree of frcedoiii modulatin- -ilit. ,tndcoclin-svstciii;thcyare,ilsojam2i --rcsist, The import,@uice of such an undcnv,-tter pattern recognition system ca=ot be over- cmpi-L,tsized. The simulation and modeliii- of it can be achieved once the parameteis of the different sensory receivers lianre been estallishcd, physical analogs derined, and models den-ised. 2 By and I -ar-e, the communication and codiii- system of electric fishes has been 26 !tbilit3, of the system has been established and discussed . The aiiiijanimin- cap, 27 -oreceptors of Stern.--irchtis all)ifrons, demonstrated through simulation The electi a fresh water weak- electric fish, have been studied, and for two different Idnds, 1)hvsical analogs have been proposed. L,cc-,tuse of our previous findinos that the anaestlictic IIAIS 222 (Fhiquel)" used by r.iost researchers of electric fishes affec3ts the frequency of the impulses emitted by the electric organs in a nonuniform way, we investi-,-,.ted a number of different -in.icstlietics and found one wl-dch does not affect the frequency of the impulses or tticir uiiplitude. 11. INSTRUI\IENTATION The recordings of the autorliytmic electric signals from electroreceptors need special instrumentation. The sigmals are of low voltages: less than 1 mv, and they need a large band,.vidtli. The diameters of some of the electroreceptors are of the order of microns. There is the need for an insulate2d microelectrode with a tip of one micron, with a reasonable low resistance, possibly less than one megohm, responding from DC to several 1,.Hz with low distortion and enough rugged to with- stand some bumpin- by the fish. It has also to be of a nonpolarizable type. We developed a microelectrode with all these characteristics and it is described later in this chapter 2 in our proposal for the continuation of the investi-ation of electric fish we mentioned 0 that we developed a sensitive low noise solid-state microelectrode d-c amplifier or 0 0 (Fi,-ures 4, 5). We modified them havin- now a Motorola MC 1531 input sta2oe and a Motorola MC 1431 output sta,- Graphs show the repetition rate versus amplifi- ,e. cation factor for square wave and frequency versus amplification factor for sinusoidal waveforms with a 100 kilooliins, 1 megohms, and 5 me-ohms input resistance (Fi,ures 6, 7). We give a descri2ption 6f the motliod used to produce a suil-ible microelectrode for recordina electric signals from the electroreceptors of Stel-ii,-irchus -111:)ifrolis. 29 lltibe?@cscribed how to mal@e co.,ttcd tuncrsten microolectrodcs. Wohlbarsht, et a2l, 10 31 (lescribed glass insulated platinum microclectrodes; Green -Lncl Grundfest, et al tlscd stainless steel electrodes. The steel and ttmgsten'elcctrodes li.,tnc -t fairly Mcrh resist.uice (20 to 100 iiic-ohms2) ,incl -,ill, iliclu(Iiii- the pl.,tt.inuiii electrode, arc pol.-irizable, 32 I)onaltlsoii descril)cs ,i inultitu(Ic of iiiieroclectrodes such .is silvcr-silvez, chloride, Ill-Itilluili-platinuiii cliloride, -aid otlicrs. Silver-silver chlorides are8 very convciiie-iit 12 igure 4. I@licroelectrode amplifier with one MC 1531 and one MC 1431 F operational amplifiers. IOA K yjl_ o2urptir IC 1,VFLIT 3.9 k CAIVACITY ydc f.AfD Pi,,-,ur-e 5. Microclectrode amplifier with one MC 1531 and one l@IC 1431 )perational amplifi@rs: schematic di-,i-rai-n. 13 Ir L t t 2 En tn -t-- T' L7- T-' i 4z I @l 2 17 '-:7: f@17 tz@:- 7 t- 7 U.Z aq 2 ta L.Li 7- ta 2 7. -7:7 -rn ::ZZ,- 7F- -77 .-'7t7 . :7- -,77M,7 2 '7 -!7 7- -1 -4 2 L.:F r 17- -.i 7 7 NV 11,1,lj@N SaAVi@l -31IVfIZ)g 'JO E-1,LVU,1401JI.L@1,Ll'; U 0.141SV- 2 kli ssoli AxoriLnoiiLiAiu r rT-, 2 T4 I 7L 2 4'' I P, It 2 I'; I T 44 Tlii I 2 .1 Tli ti 17; 2 6xr do; r,-, --T 2 0. x .).3 11).l li@ 1! I-, 14 9 rl-T" T T -rT electrodes but have a high resistance. In order to lower flie resistance, it is ,Idvisable to cover the silver-silver chloride electrode after the electrode is in.SUI,,ttcd with insulex lacquer, with platinum black through an electrolytic process xid after this to add a new silver-chloride co,-Lting. The tip of the elec2trode can be an5,%vliere between 0. 5 and 1 micron in diameter. T.,de a 50 ml beal@er and fill it with cone. H Cl and cover it with Xylene (about Use a carbon rod for on e electrode (spectroscopy carbon rods are suitable) -ind cormect it to a variable source of A. C. current from a variac and a bell-type Connect2 the low voltao, leads to an A. C. voltmeter. t@ansformer (app. 7 to 12 v). ,e LTSO about 5 to 6 volts to sharpen the silver wire #36 gauge (any necessary length) by rnovincr it up and down for about one minute (Figure 8). Decrease the voltage to lict%veen 2-3 volts and move the electrode rapidly for 30 seconds up and2 down. Have 5o ml beakers filled with: (1) sattirated sodium carbonate solution (Na2co 3 (2) acetic acid :I% in H20 (CH3CO) 3- 0, (3) Ethyl-alcoliol 200 proof, (4) Xylene. .@love electrode after shai-penincr process from No. 1 through No. 2, 3, and 4, acritatino- ..1 few times the electrode in t2he liquid. Checl@ under microscope with a microfilar ior sIL-Lrpness; if not sharp enou-h, operation 11 (2-3 volts) and.tli6 cleaning from 1 to 4 should be repeated. If sharp enough, tal@e a 50 ml beaker and fill it with Na Cl 1% solution in distilled 11 0. Use a silver wire (No. 18 to 22 -au-C) as an electrod2e (cathode) and Connect it %@-itli a D. C. source (power supply) of bet-%veen 1 to 2 volts. The positive end should I)c connected to the microclectrode. Hold it for 30 seconds in the H Cl solution. ltcn-orse twice the pol.,tri4, for the same amount of time (r, igure 9). Wash the c]C2ctrode in distilled water for two minutes. Insulate the electrode with insulex 1'.Icquer. For di-yiii-, set the niicroclectrodcs with the tip up. Di-Y for 24 hours. ri Ilic' tip should ])C clean of laccluer for 10 to 30 microns. Take a 50 ml beaker and !-I1I it with a 1% cliloropl,-ttiiiic acid. Use a No. 18 or 22 (rati- ,o wire as -i cathode AMA- A! Figure 8. Device for sh,,irpenin-- metal electrodes. i,c De\,i(-c for 'sil\-cr (-Iiloi-i(le oii the tip nii(I stirf;icc 4 of silver or silver cliloi-i(le PI."tiiiizecl cle(.-tro(les. 17 -.0nnected (to the negative) to a D.C. source of 15 volts. The microelectrode should connected to an anode (positive) of 15 volts, in series with a I Me-ohm (1 W) .Csistor. Pass current through the electrodes for 15 fo 30 seconds. N@lash the 'lectrode in distilled water for a few minutes. Use the 1% Na Cl solution with a 2 ;ilvcr wire, gaurre No. 18 to 22, as cathode, and the 15 volts D. C. source in -eries with the 1 Ale-ohm resistor for depositincr a silver-chloride coating on the :i of the microelectrode. 7\venty to 30 seconds will be sufficient. If bubbles come p Iff from any other part than the tip, it means the insulation is not (rood and should be ,edone. Wash the 2electrode in distilled water for 10 to 15 minutes. Store it in dark :oitainer filled with Ringer solution. The electrode has low resistance (from 100k to 8001c depending on the tip), is non- -)olarizable and produces very little distortion from D.C. to fairly high frequency 4$ (Over 1000 Hz) (Figures 10, 11). in Figures 11A, 11B7, 11C, and 11D, the performance of the microelectrocle amplifiers shonvn. IS 7. i 'Potcllti0iiietric device -,iiid Fi gti re I 0. resist.-ince sul)stitution ,in(I series -in, micro- 2 resist,-ulcc bo-, for nicasui electrode resistince. OFF j Fir- ,tire 11. De-,,ice for nicisurin, niicro Lid of the electrode resistance with the 5 .4 polentioiiictric device istof, mcgohm, I input r@s' - ------------T me-ohms' 5 2 ---------- - - - 10K cv)c. A-ii-,,Iificit.toti f,,ictor vp,.@su.-, rtition r,.Itc or sqtiire xv-tvc. Fl,zure 11A (2 stt-- TAIC 1 5"1 -imi)lif.) 'A -ohm inp re.% s -o @r-: me me(yohms .......... 100 Klfz 10 Kliz ,I(jo!i flet@ol vct,stt.4 freqtcrcv: sintisol@l-il (2 ci t, --c ill C I 51 lultlo,iatu al(itiol) sot3uonb iii(Itil .10[Jll(ITLir, 01)0.11051020.1.')ILU -D.Ij liplostluts pur, solici UOIIIIO(13.1 DAI',%% a.iriibs lu3.t-@)jj!p 1,c s')ittp.to3c)ll 0-TI -2li 41 2 IV, -Aa, rA C)IICIS DICITIOP ;0111 til!"t% Sil,)Uanbaij Indut joijllduir. Dpo.,103130.13TTu c).x-cnbs Itioiajjll) II, S'LITP,103z)lj (:I replosilul , S,11'Cl U .s pur '7- IL: 2 ik A SEAIZCI-L r- OR AN ELE, CTIIIC r- ISI-I ANAESTHETIC ULD NOT AFFECT TIIEIR ELE, CTRIC SIGNAL TIIAT WO reth.-tn (ethyl A commonly used macstlictic for fishes and cold-blooded animals, u 2 33 alnate = C 11 NO has been found to have carcino- nic properties by Wood carb 3 7 3 and Ball and CoNven An editorial comment accomp,,tnyin- Wood's report indicated t the substance named MS-222 (today called Finquel) nii(- be a suitable su2bstitute tha oht for ureflian. MS-222 (tricalne metlianestilfonate = C H NO S) was discovered by 10 15 5 -ds search for a reliable synthetic substitlite for cocaine (benzoil- Sandoz during I 2 35 rnethyleco-ol,-ine = CH NO ) for a local anaesthetic. 11 21 4 since Wood's and Ball and Cowen's reports, MS-222 has become routine for anaestlic- ation of fishes to facilitate the handling of both marine and fresh water species. tiz The to@dcity of the dru- to some species of fish was detern-d2ned by Marldn- 36 NVall,:e r 37 =d Schoettrrer measured its residues in various tissues of salmonicles and its efficacity has been investi-ated by Schoettaer and Julin 38 . The chen-Lical and aiiaes- 39 thetic qualities of MS-222 are mentioned by Klontz , and details abotit the chemistry 40 can be found in the l@lerck Ind= In the last.years (1970-71), an agreement has been made betxvecn the Sandoz Pharma- ceuticals, Division of Sandoz, Inc., Basel, Switzerland, and Hanover, N.J., USA, and the Ayerst Laboratories, New York, N. Y., that the latter should prodlice and s cll 2 the l@,IS-222 (tricaiiie metlimicsulfoliate) in the United States Lmder their own brand n=e of "FINQUEUI. In most of the studies on electric fishes, AIS-222 has been used -.is a general mtes- 41 42 thetic by'the investigators. Sz-.ibo Szabo and En-or , E2n-or and -44 Szabo--, Nobuo Sug.,L and m-.py others used l@IS-222 to aiiacstlictize electric fishes dur'ng suigic--al 1)rocedures and stibsequently to record the electrical activity either from the clectroreceptors or from the nerves coiuiectin- tlicm Nvitli the central No mention 1-uLs been =,L(le Of the effects t)f the I%IS-222 on the fre- liervolls syste ation and/or time. -1,ical Silr s as related to concentr Uciir 0 2 ,y and amplitude of tle elect ml 45 lock mentioned til-Lt H@rpopomtis occidenL,@lis, a South American fresh water weal-. Bul ge of 25 to'90 per second, would lower jectric fish, with a normal repetition rate ran C.- 2 sia. Under deep anaesthesia, to below 16 per second only i4nder anaestlie its frequency the fish may stop abruptly its electrical activity. By reducing the level of anaesthesia al r by stoppin- it to let the fish recover, 2it will also abruptly resume the norm electric activity- In our ex-Ln--riments with Sternarchtis alljifrons, a South American weal.: electric fish, we found a gradual taperiiicr of the repetition rate of fresh water the electric signal with the deepness of the anaesthesia and a gradual increase of the rel)etition rate N@,itli the recovery. B2y monitoring the effect of Finquel (T@IS-222) amesthetic on tl-ie electrical activity of Sternarchtis all)ifrons, it has been observed a fast and sianificant change in the rel@etition rate of the electric signals. A decision %%,as made to study the effect of different anaesthetics on the electrical activity of different species of electric fishes. The anaestlietic acren2ts of choice were: 1. MS-222 RINQUEL TRICAII\TE METHANESULR, OLNATE 2. NE',NIBUTAL 3. A',% IY T A L 4. SECONAL 5. THIOPENTAL SODIUM 6. NOVOCAII,,TE 7. TERTIARY AI%IYL ALCOHOL A. MISTRIJ'.NIE'.NT2-fATTON, 'AIATERTALS, AND -.,@IETTIODS A sl)cclally built "1)0 cm tr,-ty made of lucite which could be acljtiste(I to t'lle size of the fish ILis been used to check the effects of ,tiL-testlictics on the electric or--.ins of clcc- tric fish (F!1,.,Ltrc 12). The tr,.ty is I)rovided,%vitli fittii-i-s for the r.-,pid disch,-Lrge of 25 - ---- ------- Lnd a constant -Icration of the solution is possible if necessary. ,Lestllciics or water It also has at every ceiitimetcr distance ciiil)cddcd stainless steel electrodes con- Iectcd on both sides of the tray with the exterior and to contacts. The electrodes 2 corresponding to the position of the head and tail of the fish were connected through 2 W-Th--,-Lin, low noise amplifier to t-%vo oscilloscopes Telctroilir. (one for plioto-taldiig), a counter and .tii FAI tape recorder (Figure 13). The tray with the fish was located in floatincr screen room and all the instrumentation grounded to the scr2een room was located in the l.-LI)oratoi-y outsidc the screened room. A lucite cover on the tray pro- vented the fish from jumpincr out, different holes served to pour in water or anaes- theticsto put in tliermometers.,and to let excess air out. A d-c (battery) operated high iutensity lan-il) was used to illumin,,tte from the top the fish and to fu2nlish enough heat to hold the temperature constant during experiments. For the experin-lents, the fishes' own aquarium-water was used at the begilinin- of the experiment and the same water ,Nas used for nii@dii- in the anaestlictics. In this way, the temperature of the water and anaestlietic solution was easy to keep to the same level as the normal temperature of the water in the aquarium. The pH of the aquarium water -,it the be- ginnin,,,r of tl-le experiments and also the pH of the -maestlietic solution were measured with an ex-paiided precision type of pH meter. The pH meter has been calibrated before each e%-periment. Tentative measurements and observation of the action of the anaest2lietic ha-%Ye been made first oncroldfish and then on one of the electric fishes before another specimen has been selected for the experiment. For each experiment with one and the same amestlictic, five specimens of Stern.-irclius -tll)ifrons have been used. The specimens of Stei-ii-,Lrcbiis -,ill)ifro2ns, a fresh water South American sternarel-iid weak electric fish, have been in our laboratory for over one ye.-tr and they were all healthy 'Md V-"ried in NNci,-Iit from 14 to 30 grams. G@,i-nii.,Irclius niloticus, ,t fresh water African gyninarcliid wc,-d@ electric fisli,'lias been l@cl6)t for over six months in our l-,Iboratory. There is -tnotlier specimen just received. There ,Ire also two Gnttlioncmus lvtcrsii, fresh wiwr African niori-n)-rid elect-i-ic fish, bcin- in the lal)oratory for over two nioiiths. 2G '7" -7 re :L2. Adjustable Lucite Tray for An,,Les tliesia Experiments. Fi-tirc la@. Instriiiiiciitation Used for An-testlicsia Experiments. 0 27 riment and put in the experimental The fishes have been weicrhad before each expe ium water. The electrical activity of the electric oro-an has tray in their own aquar ded wifh the FM magnetic tape recorder, its amplitude meas - been monitored, recor calibr2ated osciuoscope, and a photo talcen. After a few minutes, the ured bn the -water was discarded from the experimental tray and the anaesthetic was introduced With a funnel through one of the holes in the tray cover. The time of introducing the anaestlictic has been marlred,%vith the aid of a timer and also recorded on the magnetic tape and in our records. T2he effect of the anaesthetic on the fish, its behavior and reSpirELtion were constantly observed. The electric activity has been monitored and from time to time a photo has been tal@;,en from the oscilloscope. The moment in which the fish was anaesthetized completely has been recorded. The fish respira- tion and electric activity (amplitude, wave form and repetition rate) were constantly 2 observed. If necessary, the anaestlietic has been immediately discarded and fresh aquarium water has been introduced in the experimeatal tray with adequate aeration for the fish. This moment has been recorded and the recovery time of the fish has been marl@ied. The electric activity also has been monitored.. When the fish was considered completely recovered, it was returned to2 its own aquarium. Sometimes because of the long returned from tl-ie ex- time a fish was anaestlietized, it has been parimental tray in a net floatin- in its own aqiiarium ,vitli adequate aeration ,vitli bubble stones under the net. Expe riments were pdrforiiied on five specimens of Stern2archtis ,Llbifrons (No. 2, 3,4, 6 and 7). In order to assess the effectiveness and dosa-e of the "MS 222 Finquel" tricaine metli,-mesulfonate, we recorded the electrical activity before, (.Iurin,-,, ,tnd after anaestliesia. The frequency of the electric org,-m will drop iminecu,-itely after 2 addbia the tric.-une mctl-Luicstilforate to the water in a special trayprovided with stainless steel electrodes set at a distmice of one cm from one another over the lcngth of the lucite tray. The fish were restricted by a p.-irtitioii and a "Ull shaped lucite (IcN-ice pl-,tccd on top of the fish. Fi-ure 1,1 sliows the decrc.,tsc iii frc(Itioncy 28 t 11-171 @4 i-I FA) 2 1- 1 71.::i t -tit 2 V-4 r-4 2 C3 rn 0 2 en C) 2 CD Clt ::i 0 2 rp C4 7@t-7, 4 2 CD C) C) C) c@ CD C> (D CD CD (Z Lf) C4 C> (O tD (D uoT slr 1 'du,, Ir sa )Z)1:4 10 ol solution in n the fish has been kept in a 1:10000 MS222 Lesthes ,,vitli time,%vhe alL ia wawr and the return of the impulse rate after the solution has been exchanged with ittin2g organ was recorded fresh water. The electrical activity of the electric transm on a Hewlett-Paclmrd four-channel nl magnetic tape recorder to be played back and analyzed later. Figure 15 shows the electric activity before anaesiliesia; Figures 16, 117, 18 and 19 show the electric activity during anesthesia; and Figures 20 and 21 shonv activity during the r2ecuperating process. C'USSION From seven anaestlictic agents we tried, six were inadequate because the repetition ate of the electric signal was affected. Only the thi ental sodium Abbot (pentothal r OP sodium) does not influelice the electric activity of Sternarclius albifrons. The tliiopental sodium has been checke2d on five different fishes (same species). It has a very fast effect in a dosage of 1:10000 in,%vater. The fish, after it has been a=sthetized, %@ould remain ,is such for 2 to 4 hours lyin- very quietly. It recovers completely and the anaesthetic has no ill effect on tl-le fish. We anaesthetized them repeatedly and after six months' time they are doing wel2l. After the fish has been amestlietiz,ed, it has to be put in aerated fresh water where it can remain for hours. Figures 23, 24, 25 and ;6 show that the repetition rate of the electric signal has not been affected in over 40 n-dnutes. The only cl=,-,e was produced by a slight de- crease in water temperature (from 720 pulses to 700 pulses) of al-)out 0. 4 C. 2 Sternarchus albifrois is very sensitive to chan-es in water temperature. It will increase the repetition rate of the electric signal for an increase in wate7r temperature and it will dccroase the repetition rate for -. decrease in water temperature. For every degree Centi--rade, it may chaii-e the repetition rate by about 50 to 80, de- Pendin- 2on the particular fish. TluopenW E@odi,,ni or SocUui-n 5-ctliyl-5-(I-mctliylbtityl)-2-tliiol)arbitxirate has the chemical formula C H NaN 0 S, a molecul.-Lr wei-.Ilt or 2G4.33 aiid is a yellowish- 11 17 2 2 L 7 St,,Illl,c,litiq ,ill)ifrons T'r7 normal elee- Fig.16. Sternarchus albifrons #li in anes- -11 -.Icti'llity - Water temp. 24. 2'C, time: thetic tricaine methan2e sulfonate 1:100-00. cc I msee/cm, /cm, niV/cm.. Water temp. 24.2'C, time: 5 mV/cm, 1 minute. .It rig ill)ifi-on-, 7 in ,iiie s - Fig.18. Stern,,ir(-Iitis ill)ifi-oiis !.'7 in -,iiies- iiietliiiie slil foll.-Ito 1 :10000. tlictic t-.1@iciiiiL% iiietli-,iiie sulfoiiate 1:10000. 2'C8, tiiiie: 1 iiisce/cm, Watcr tc)iij). 2.1.2'C, tiiiic: 1 i-nsec/ciii, 5 iiiV/cni, 4 iiiiiitttt!s. Fig. 2 0. Sternarcliti,,; -Llbifrons recuper,,itin", SteriiirclitiS -tll)ifrons ;@':7 in anes- tD tric-,iine metliane sulfonate 1:10000. in fresli water. Temp. 24.2"C, 5 m2inutes. temp. 24. 2'C, time: I msec/cm, -IV/cm, 5 minutes. -7@ L L,,:7 It 'AVE SLLriiireliti-, recul)cr.,itili.- iii r- ig. 2 2.. New sct-ul) for microclectro(le %v.'tter. Teiiip. 2.1. 2'C, 8 iiiiliLit(,-s. recor(liji,-. 1"7, L Figure 23. Sternarch-Lis albifrons- No. I just before anaesthesia with tliiopeiiial sodium. Electric sirnial rate: --710. SNveep 1 ms/cm, Gain 10 mi?,/cm; Water Temp. 22.4"C. Figure St1ei-iiarelius -ill)ifrons No. I aiiacstlictized with tiliopolital sodium 1:10000 (2 n-dii.) 3.5 min. in fresh w,,ttcr but still coiipletely imiiiobile. Electric si,",Dal rate: --705. I ms/cm, Gain lb mi,/cm; IV,,ttcr Temp. 22.35'C. Finure 25. Stci-ii,,trclius albifi-ons No. 1 amostlictized with tl-dopental socliuin 1:10000 (13.5 min.). Electric signal rate: 700. Sweep 1 ms/cm, Gain 10 niv/cm, Water Teml3. 22.3'C. 71, Fi,-ure 2 6. Stci4ntrclius -tll)ifrons No. 1 "aesthetized,%Niili tliiol)cnt-,il so,,Iitiiii I. -10000. -07.5 iiiiii. in fresli water. Electric signal rate: -@j 700. Sweep 1 ms/cm, Gain 10 mv/cm, Water Tei-np: 22.3cC. - -- - --------- ,vliite, hygroscopic power, soluble in water .L4id alcohol -Uid is a strongly alkaline 46 solution. The Abbot preparation is of nonhygroscopic crystals Tub0curarinc cl-doride can be administered by intra-al:)domin',I2l injection. There is no interference bct%%,ecn tubocurarine alid tliiopental sodium. The stability of tliiolp-ntal solutions depends upon several factors, including the diluent and conditions of storage. It is recommended to keep them under refrigera- tioii and tiolitly stopl3dred. ,T.mquel" or IIAIS22211 is a meta-amino-benzoic-acid-ethyl-este2r in the form of tri- caine metliane-sulfonate and has the chemical formula C10H 15 NO5S with a molecular weight of 261. 31 and is produced as fine needles, soluble in water - It is slightly acid and is stable to boiling. Finquel 1:10000 in aqliarium water would affect the repetition rate of Sternarchtis albifrons and make it decrease in 10 minutes from 780 to 440 (Figures 27 and 258). 35 Fi,-Ure 27. SternarclliL-ls-a-411-II-)i'fLr@IOJ-ll l@ @s-02 in Aquarium Water. Rate: 780. Electric Si.-nal Rerietition 22.4-C. 8/31/72. Am-plitucle 25 mv, snveel:) 1 Ms/cm, -%vater temp -'Pik ;7 Fin-Llrc 2S. Stci-ii-irelitis -,tll)ifroiis No. 2 in T%IS-222 (Filicitiel) Al-Licstlictic 2 oil ILitc Soixiiion 1:10000 ill W',Itcr. Electric Si,-,iL-11 Rci3ctiti 4,1 0 . After 10 in tllc All,'Lestlictic Anil3litude 38 sNN-cel0) freq. I n-isec/cm, Nv-,ttcr tcml). 29-.,I'C, -S/31/72. IV. TECHNICAL DISCUSSION Four specimens of Stcrnirchtis -,tll)ifrons have been used for esltblisliin- methods to record the autorhythiiiie activity of their clectroreceptors (Figures 29, 30). The r,shes hane to be an,,testlictized and cul-arized in order to avoid twitching of the muscles during microc2lectro(le recordings. We tried the effects of d-tubocurarine on 42 stl,rnarchus ilbif i-oiis.. Ha,,-,iwara, et al , recommended 0. 05 to 0. 1 m- curare/fish 43 and Enger and Szal)o recommended 0. 03 mg d-tubocurarine/,-, fresh fish weight. I3,Dth used (,@IS 2222) tricaine methanesulfonate for anaesthesia (1:150, 000). We folmd that the quantities given did not correspond in our case. The weiaht of the Stei-narchus specimens varied between 16 g an@d 19 g. One specimen (16 9) recei-,,ed 0. 5 m- d-tubocurarine intra-abdominally. The fish was paralyzed in one minute. Electrical acti,,,ity of the main electric organ subsided after 20 minutes and the fish was dead after another ten minutes. All the time the fish was kept in a 4000 ml beaker with medicated aquarium water at a pH of 7. 0 and a temperature of 23.OOC. An aerator stone provided the necessary air. A second specimen of Sternarchus (also 16 g) has been anaestlietized with MS 222, 1:25,000. one gram of MS 222 has been dissol2ved in 1000 ml distilled H 2 0 as 'his was diluated to the proper amoun in t a 4000 ml be.-tker by stock solution. Then t adding P-quariuni water with merbromine and acriflavine added as disinfectants. The MS 222 dilution of 1:150, 000 would not affect the fish in over one hour. The dilution of t2he '@IS 222 was reduced to 1:25, 000. After 20 minutes in the MS 222 solution the fish was injected intra-,.ibdominally with 0. 05 mr of d-tubocurarine I mil solution. The fish was paralyzeci in three minutes. For one hour it gave a good stron- electric si,-m r i.-il of the niai6i@ clecti-ic org,.ui, aft@--r. this cr idu,,Llly it diminished in stren-th and in another liotir the fish was dead. Cb Fi(Turc 29. Set-tip for recordincr the autorliytlimic activity of electrorcceptors. Ile Z L Closc-lip or illc, s(,t-ul) roi, rc,(,or(iiill- ille Fi-ui-c 30 of clecti-ol-(.Ccl)tol,s. s,ponci for our .drlot corre b-y the Previous autilo rs di .,j the ,neus - stlletized ia MS 222 2 8 %) ,Vas first ana-e ith (.1 ml of Stcruarcl-lus ted intra-a'3don'lina-11-Y '%v - third SI placed then a "is it O's ini2c' for 20 Tninutes 31tcr tl fish "as bee rarine (r. igurc 31) - The &cri- d-tubocu , acroLW cl ,,.,aler 'I@lith rnerbromine 1s beer, it'h circulati the electroreceptors Ila tray -rom ,,ial reco (00. fler-or(lincor f silver-silver chloride .1@,IS -?22, 1:'7@@01 in t - r ,tnd a Ville and ,he tra,, 'ic,,,re 32)- T"e th a iieutr,,l electrode electroreceptor ,,,,Ipted'wl placed OrL a tonic I Ifipullar5 ttiag electric Organ 3.slred the -,vealr, -croelectrode -a' ed to the inalu from trans' -,pecilneu has been return "is s strong 2 electric of the ele,troreceptors. T' ,vell since then acti""I s and recovered alrnost instantly and is doillo- ter 3( Ir.Illute treatine-at 9-1 the 2 g) received the sa,,ne -- ,vster, in tllc tray imen Of electrode .knother lourtl-l spec. stead of using the neutr2 0-aic, cimen. TI-as tin-,e in jallieter %,%Yas placed around tle t signal third spe ride %vire loop "/4 in. lectric Organ silver-silver cl'10 2ic-rure 33) - Tlis tirae the -main e electroreceptor ,mpullary electrorecel)tor ("' - activity of the "S of the authorh'Yth'n"c recovered aln-iost 'nst3-ntly' t Inaslr 2 to its tanl,, and (-fia did no the recordin'. -liraed since ti-ien Urc tl, , ill effects 4o rnin'Lltcs I e fish NNas re2, io!Llt an@ .kiter 0 @-peciine", qnd is do']act fine la-,e the Pre-,-iOus (I rooln frotn 2 i'a 34). 'I sldclcle e,ts "v er e pc,_ roo"n cliiicor Lte aiid The Tnicroelectro(le recor corlls Nvere elilaill, ca xvas done by a nlicro- ts, I)Iucrs In(, struine'a 0 2 -1, f 1-,, speciril ,wmch M-c Illunlil'Ition 0 ,,It car baticry' All stit. co -a-lectca to ,Itccl , 2 'L no,,it s co,, -ilicroscollo to bnttcr,e t,^ C COIUI 2 tllc @rco-I itiers N,,cr 0 jaclr fil-l@-Cli 01" s the 2 of thc,%,votcr Olit oscol@,c sul)port, tlle n.,icroscOI)cl -1-licr ,ts cnt, SLl ell 6CY,her )ifroll'3 S13ecirnen after ,c Ficrur II. i- i-n einject@i(o d-tubOclr3L'71 tollic elcct'7 recc 9 I)tors. 32. . )Oil @-, -ire I-C c@c I)LO r'- - LI )@l r-I -C ttillcl.o"g, 4( Ira t r ilk- activity of alnpl"13@ry for rcc0l:dln'T auto all)itrons. The silver- 'r,icrure Set -L r clill 2 is visible - e ctrorecePtOrs of Ste'rll' I clectrocl ele p neutra ,,vire 100 -c silver Iiloride tile 2 tell,, nll ctiv 1 ty C-irl)oll 34 tl,C oscilloseol,)C" reco tr, 9 ,Iittil A-1 ,plifiers, etc., was connected to one point ground and tcL the Tektronix T22 modified Lferential amplifier ground. The rnicroolectrode amplifier and support was changed Om the intcotated circuit model used before to an electrometer tube type and a three -dch is less sei-Lsitive to the change in impedance produced by .alls2istor amplifier Nv.1 ,Ii@,lit movements of the fish than the previous one. The microelectrode, a silver- siln er-cliloride, platinum, silver-cliloride electrode of 0. 5 micron tip diameter, was positioned on the electric receptor inside the groundina, loop. r, igure 22 shows the 0 recordina, setup and Figure 35 t@e sche2matic of the microelectrode amplifier. At the same time, the electric activit@,- of the receptors was displayed on one of the traces of a dual scope. The activity of the electric transmitting organ was recorded V,ith two carbon electrodes placed at the end of the tray and connected to another TektronLN T22 amplifier and displayed as a second trace on the scope. A Hewlett- pdclmrd four channel rM tape recor2der was used to record the electrical activity of the electric rc-ceptors on a inaoiietic tape. It can be played back and analyzed at a later time. Photos Nvere made dui@tig the recording. Figure 37 shows the activity of a s@ncluonous tonic electroreceptor (in a previous report we mentioned that we re- . corded from a nonsyiichronous tonic electroreceptor), and FigLir2e 38 shows the elec- tric actiiit@, of a nonsynchroiious phasic electroreceptor. After finisliiii,- the e.%I)eri- 6 ment, the fish was injected with neostigniine metl-lylsulfate 1:10 to counteract the effect of the D-tubocur-,triiie. The fishes recuperated in a few minutes and are doin2cr well. Another c.,,I)erinient was performed with two Cr@Zn-ui,.trclitis ililoticus, Cuv. Each of these fislies was placed in 25 ff.-dloli water tai@@s with lucite trays raised to ,tppro.%-i- mately 4 inches from the tol). G@,iiin-irelitis niloticus is an air breat2her -tnd if left in a deep tard@c mtist c.%I)ciid execssi,,,c ener,,,,), to swim to the surf.-ice for brcatl-dncr. 'nic tax& is kept clean by lucite plates provided with lioles to let (lirt fall to the bottom where it can- be N-.icuuiii-clc,-uictl very cas!13,, tlirou-ii speci9@a lioles. The t@i@@s of 2.5 V@f 2 30 c o,fiTP13'. 2 2 5 PL r-4 C41 ZPTJT 2 GPD ColniBCTO'R 7 6 .'V IL. 5 C4 2 GA114 0. 01. 0. ol i@f r 'V + 8 + 2. ra 'V 3.6 :Sze ,@d, c3thd. to wtigh unpgdaWC mi'T.wiect Syn- -Fig. steriiare. i,,:oelcctro(l2e, clir loo Inv/cln. Ms e recordin<-r ilnpellaiice -rO 2 elect -ore le iollo"ve-.. ir -qv f 7.:,! tA 72-i 39.- of tilc tr.'U'l -fie 0 tile 0 elect 2 rlll. colalec s or4flul. vot T@licro- electric r C'11" "tl.ol-c(,Cl)tol" 0 Iiie iliv/cl-il. or mloiller iiisce nn,,trclitis. nilotictis are approximately three feet apart. The covers and outside ,niagnetic stainless steel frames were all grounded and so was the taijit through .bon elect;i-odes placed in the filters. @o carbon electrodes in lucite tubes (Nvith holes) Nve re placed at a distance of 15 cm )m each other alongside the fishes. A third electrode was place2d either midway ,t@%,een the recording electrodes or on one of the ends of these electrodes. The first .,o electrodes for each fish were connected with the T22 Tektronix amplifiers and )nnected to two different Tektronix scopes. The third electrodes were connected ) a DPDT switch which could actuate also a battery to raise the lower trace of the cope,%vhen the two electrodes in the t,.vo tar@cs were connected and hence a commw-iica- ion link- was established bei-%veen the two GNmnarchus, to study the effect of social nteraction and communication bet,@,een them. n Figure 39, the normal electrical activity of the electric organs of Gymna7echus T ,o. 2 and in Figure 40 the electrical activity of Gyn-u-iarchus No. 23 can be seen. ,!(.rures 41 and 42 show that Gymmrchtis No. 2 almost stopped the electrical activity )r ten to t%venty seconds. Figures 43 and 44 shoiv- the modified activity of the im- .Jses from Gynu=cluis No. 3, and Figure 45 shows the modified activity of Gyn-ui,,trchtis o. 2. When a metal rod was introduced in the tank of Gymn,,trchtis No. 3, G32rrnmrchus 2 would nervously move back and forth and eventually attack the electrode connected tli the other fish. 'niis is only the beginnin(r of studyin, the commlulication 1-,---t%veen o g=archus niloticus. a gnn-nnarclius niloticus baby was sectioned and f:L%cd in buffe2red formaldehyde (10@o) about one month. Then it was dec:acified with Kristensen's decalcif),in- solutions 24 hours. After tliis, it was deh3,&-,,ited usin- ctliyl iilcoliol, toluene, toluene with ,tffin, mid finally eml)cddcd in de-. ssed p.-tr,-trfiii with 56'C iiicltiii- 6point. 'Is CLlv. @r2. irchus nilotlcl -Fig. 41. GYnln-, n -lectr C o2rcran wllell S cliv. 0 Electric a "'roucrh Lirchils ililoticli Iectric3.lly connecte'l ' 0. GvlnT 2 the trails electric ta:a@, has been . (I ,L wire and S,*,,?itch%N'Ith ,tri d with the Of anotlier carbon clectrode5 an .-"L's 2 mscc/cr-"; 3n. Not conno ti-ler G-,,i tie tarar, of ano n-isec/cm; 1 M'%7 CUl /cm. Inv 7.' !7'i2 7f Z7 ic,Lls Cllv. ,ricr. 43. rul NN'llen 2 17c- o@r CD Elee- ty of clectri ctc. Electric '.Icti -illy coilllc LI tllr 112.2 c@ 'C,, C 't2. 11 t IC tllc t, Icctric lIcLs b 2 es 'u,cl electric orc@ , 11 t@,,l I @t lvire let ic_ aleetc(i li,rc C,,tl:boll clect-j--'cK' 2 "Isec/cl": t Illy co,, 'u", SVitell \Vitli a CICC -v C tile tiu @Or ,Illotllcl: bo clcctro(Ir-s U.I(l -,L \Vir 2 "iiiscc/cl"5; ,,v/ci of , . G-\-ilill"r("lu- ,uiotllc@ ,,V/cni. 1--7 @W E r r -T,2 F -iriirchus niloticus Cuv. #3. irclitis nilot2ic-Lis Cuv. 44. GNrn Fig. 45. G@rmi, 7; 3. -ject.ric activity of the electric oi,-,,,,m ivlien EleetTic activity of the electric organ,%vhcn targc has been electrically connected the tar2@ has been electrically connected -odes and a2 %Nlre and tlirou(Til carbon electrodes and a wire and .rough carbon clecti 0 -;itch with the t,,uilc of m-iotlier Girrri-Lircl-itis. snvitcli,,vitli the tarfl-. of another G-,,ninareliLis. msec/cm; 1 mv/cm. 2 msec2/cm; 1 m'%I/cm. After one iniliute, the fish switched to its normal acti,%rit3r but with a smaller amplitude. An A. 0. Spencer microtoine has been used to cut 10 micron slices. They were stained with Hem,-ito.Vlin-Eosin and mount@--d with Per2moiLmt on iTAcroslicles. There are about 27 microslides with four to five serially cut transversal slices numb,-.red from the head in the direction of the tail: 1-1, 1-2, 2-1, 2-2, ..., 7-3, 7-4. A few photomicro,-raplis are shown as Figures 46, 47, 48, and 49. The interpretation of the 1-dstolocrical preparation will be made at a later date. (Ficure 50) 0 0 %Ial-,tptci-tinis elcci-ricus, the electric catfish (Figure 51) is different from the other ,loci -.tciiii's I-,tNv. All the other electric fishes Tic fishes because it does not2 obey P ;bey P,,tciiii's laN%,, according to which the innervated faces of the clectroplates be- ome ne-ative during the discliir-C, whatever the arlttdmical oricnt-Ltioi, or the orcran. 47 'his fact is due to the uiiiclue ai-Litomy of Alilipteriii-tis. Th6e electric or,-.-M of C d .alaptcrtirus forms a sort of loose jacl@et ,trouiid the fish, iiist ad of I)cilr,- onibe(Icle I'll..@, jj .., L jv L 'ITICI 0111 JO LIOTIO@-).ITP -)III If[ T-/T 4sul otil li'@'1110.1111 Ino li;olo,kst2n;.Tj, (Scl-uco oAlID Isti I g:p 1 -J .101111 SIILLD.IU 9 2 .4 t -e, 4 baby Fig. 49. 1-djoticus Cuv., baby pi1w cord. 2 molitils). Spinal cord. pliotoinicro- erferciicc 9rt-pli Nil@on2 pl,abo-interforence I\Iicroscope x 2oo. r.-L%-cd:' rormaldcliycle, 10%. De- -aalc x So. r. i.-.:e d: oi-i lel'Yclcs I t ii.,irc-,Iilis 1-tiloticliq Cii-\,. , b.-tliy 13r,iiii. I'lliotoiiiicrolvl-.IPII -iiii-erfereiice T\IicroscolY-, x rorm,-,I(lclly(ic, 1070. De- nil-.I 1111 LI E NL C9 70 5 0 ADS L 2 2-5 m.n rn ng xvlcne ALC .3 min 3 min C@ 3 min ter 2 3 min 3 min 3 min 3 min Ott S 04 1 02. .03 min 2 _in@in@ 2 r was absent Wate If mercuric cbloride ve, skip Steps 59 69 2 from fixati and 7 and proceed 4 to S. etc os n rnin 2 0 IC4. L Few Dips B B lc5 A L 2 ALC Xy n L ene Fe-,v Dips 3 min 3 min 3 min 3 min 3 min FIELD IIARRIS lir.,MATOXYLII2'4 ig. 50. DE LA thin the body muscle as in all the other electric fishes. The electric or,-,-Ln, depend- on the fish's size, forms a sheath from 2 to 10 mm or even tl-dckcr, in which the ,cti,oplatcs are st.,tel@ed in the longitudinal direction arranged in a somehow non- -larly fashion, contrary to the oilicr electric fishes. The electroplates are disks about 1 mm diameter and 20 to 40 microiis tliicl,3ioss and derived from myeloblasts. the center of the c-.itidal face of each electrol)late, a thin stalk arisesfrom a complex agiiiation. The end of this stalk makes contact Nvitli the branches of a single myclinated -ve iner arisiii- frou-i a large ncrne cell situated 2in the gray matter of the spinal cord ween the levels of the first mid second spinal nerve roots. As it can be seen on -ure 51, the nerve ti-ui-Ic which is heavily myclin-ated is- easily identified. The way ,,Iiich tiie electric or--.m is innervated plays an important role in the synchronization dsm of the discharge. 'i@lal,,tpterur2idie discharge consists of a long train of 15 to 30 monol)hasic pulses to 2 msec duration in a total time-internal of appro.-dmately 100 milliseconds. '.x to seven inch electric catfish could discharge impulses of approximately 200 V. ily surprise, Dr. Earl Harold froiii the California Academy of Sciences and Stein- It Aquarium in S,,ui Francisco, told me that the 2two very large (appro.,dnutely 2 to )ot) electric catfishes which tl-ie aquarium had did not discliar-e more than 100 volt eS. surprising that the ikl,,il-,ii3tci-iiricl,,tc we checked could.put about 0. 5 Av/lu- of energy !ach gram of elcct-i@ tissue of the electric orgaii. It is our intention to investi-,'Lte fact more closely. The electric discliar-c of three stronc, fresh water electric 0 t3l s i\l-,il,,Lptertiriis @lectricus wei,-Iii-Li- between 55-65 g was measured when in the r_v 8 r -uid out of water in the spccial tray with conticts. They disch,-trged between )O volts in ,v,,tWr mid I)ct%vecii 150-190 volts out or Nv,,itcr. The (liscliar,-c consists rsts of 5 to 7 inil)ulscs of 1: 5 to 2 nisec dtiratioii (ri,-uir-s 52, 53, 54). Figure 51 MalaPterurus glectrtcus. Th Nile Electric Catfish. (Shown.at 1.5 Scale) u Filr re 52. A-Ti l@ll')tertirtis elcctrictis. African stron. fresh water electric fish oil the scale: wei-lit 61. 5 g. co .......... Figure 53. Dc,,,-ic5(@ fol-,Inclslirillcr tile %,Oltl-c or the Cliqclllrrr of es E. k. L I-i-ure 54. Discliar-es of the clecti-ic or-an of 1,@T,-tl,--tptcrurtis electricus we lia,,,e now in the li.lioi-,ttoi-y. Vertical: I g2raduation = 50 V. Ilorizoiital: I gradu,-itioii = 5 nisec. La our s5,noptic Table No. 1, the past achievements and results of the study are men- tioned, the proposed continuation Niorlc is sho-%vn, and the f-Liture objectives are delineated. rhe multimodulation-multicocliiig communication syswm of electric fishes ser2ved as a nodel in the development of a con-iniunication teel-a-dque to resist jarnn-lin,-,. 'he physical analo-s of a phasic and a tonic eiectroreceptor been established for 0 'ic Stei-liarchus all3ifroil,3, a Soutl-i Americ-,tii fresh water we,-Jz electric fish of the I-ii2ch t7 .,Qquency t@,I)e (riniircs 55, 5G). The eleettoreccptors of other species have to be -lie rel-,itioiisliips of the various clecti-oreceptors to the ulidenvater -vesti-ated and t 2 0 @ttern recognition systeii-iiN?ill'be est,-tl.)Iislied. nientioiied the effect of nion-iii- objects p-,ist the, clectroreccptors iii-one or -uiotlier 0 ,ection. Some of tliciii Nvoul(I respond iii one w.-iy for the foi-%vard direct (head to WI) iiicrcasin- the r,,tic of the iiiil)tzlscs and in another way for the bacl,-\vard direction 'rAJ3'LE I PA$T, ACTUAL AND FUTTJRE STUDY PROCRAAI OF UNDER%VATL-R PATTERN RECOGNIL"rlObtl SYSTE,%R SE,t4SOR STUDY RE. suLTs 2 r-ItELnUNAP.'f ANALOG IVIODEIS . Asynchronous Phasic ST 0 t-TuIU-inodulatioxi 0 Asynchronous Tonle - synchroiioti2s.rinic PI[JISE I PILASE 11 a Investlg:tte clec'troi,ecelitors of other species 0 I)osl,-n & conduct s(inultis-response 0 Estai)lisli relauomliip of various electric exp(,,riment@', Jn lai-ge tank ijLIY OUI'IILIT )PO-CED recci)tors - Determine effective range c 3-- (INDL;'R'V,%TI:P. PITTI, 2 o Llc-O'Crtjiijio lio,.v receptors are used In o Sign:d processing In volsy environment RECOC.,N-1-i7ON ntvi-allon 0 Improve(] pliysleil nnalogs 2 PArl'EJ14% )VL-,(:(,)GNI',' 07liEr% SENQOIW 2 Z.IODEL inE tleclianlc,-d o Cross Coi-relation Sonic ,Nlajority L--clsion Optic i-v r r el e2 r r ec ec < rs r ee rg r 2 is G a ri]L ri2 s Figure 55. Tonic, ai-iil)ullary electroreceptor. Pliysical analo-. current ,md volta-e r generator internal resistance 9 !2 exterml resistance G a autorhytliiiiie generator exterml can,-LI resistance ril ri2 internal resistance external canal c,,Ll)acitaiice c s internal receptor cal)acitmice receptor iiuicr face resistance r s sldn resistance i-v re, re2 des r ic ic c r re 8 r 9 r is G 2 a ri, ri2 Cs Figure 56 Phasic, tul:)erous electroreceptor. Physical analo.-I. current and voltage r 9 generator interml resistance ex-terml resistance G autorhythmic generator.2 a internal canal resistance ril; ri2 internal resistance' internal canal capacitance c s internal receptor capac'it:uice receptor cell c,,tl:)acit,,tiice r sldn resistance s receptor iiuier face resistance c C.N-terli.-tl sidn cal),-icitaiice es (tail to head) by decreasing the rate of impulses. Other electroreceptors will react to higher or lower conductivity than the medium (.iquarium water) in a similar way. Obviously, the response of the electroreceptors in other electric fish species have to be assessed in order to get a better overall picture of the underwater pattern recogni- tion process. Figures 57 tlirougli 63 show two d2ifferent electroreceptors of electric fish. The results obtained from the electric receptors study will lead to a simulation plan of an underwater pattern recognition system. A passive hybrid underwater pattern recognition system bloclc diagram is shown in Figure 64. In this bloclc diagram beside the electric sensory receptors, the other types of lateralis line sensory receptors are combined to obtain a better cr2oss-correlation of the different signals. Eventually passive optical sensory receptors will be added (for detectin-, for example, changes in the bioluminescent organisms in the water when disturbed by a stimulus lilce heat, ivater turbulence, etc.). With regard to the other lateralis line organs, Sternarchus albif-rons, like other teleosts, have canal orcans as well as superficial organs, called fre2e neuromasts. Some parti- iial or-,-ans are in direct relation .ularities of these organs should be noted. Not all ca -,vith the external environment. The sense organs do not always alternate with the pores 48 -, are located between two pores. All :)f the canal. In some cases, three sense orgarl ,Iwee cupulae lie in a "watery" fluid filling the canal. The fluid seems to be a form of )olymucosucharides. The canal is not free from one pore to another, but is obstructed )etween the front and second, and the second and third cupula-sense or,-an units by'hvo @pithelial plugs Thus, the i-niddle unit is not in direct continuity with the external en- -ironment (Ref 23). It should be mentioned that Gymn,,trclius as well as Z@otoptemis @ave a completely closed canal system. The free neuromasts found in the head of teriiarchus are relatively 1,-tr,- ,e, are protected by paired prominent epidermal flaps nd have.a.histological strticture similar to 'big pit or,-,aiis". As in the pit organs of 0 @vmnarclius (Rdf. 25), these free organs of Steriiarchtis are innervated by a btindle AZ LA.. 7 ci -- - ------- CD Cil 90 En 0 2 0 rn ri 0 rn 8 rj C+ 0 ectrEP 2 (CIL$ 59. Ampullary tonic electroreceptor. Fig. 60 Stimulus and recordincr from 0 (Schematic) an ampullary toriic electroreceptor. 2 eP lits iiid recordin- from a 6 1 Tul)erotis I)hisic clecti-oreceptor. Fig. 62 SIL-ii-nu 0 (Schematic) tubcr(xis pliisic elcctroreceptor. Gi .. ..... ... I2- wt 6- c Fig. 63. Electroreceptors of Gvmnarchus niloticus. a: ampullary dlectroreceptor b: tuberous electroreceptor t3rpe b c: tuberous electroreceptor t@-pe a. (Schematic) 62 FIGURE 64 PASSIVE HYBRID UNDERWATER PATTERN RECOGNITION SYSTEM L EL EL I III z 0 ipassive Passive 2 E ME Acoustic Optical z 0 ]I Rec. Rec.. c@d P-4 Displ. Dispi I II EL I to IV = ELECTRIC SENSORS 5 ME I, ME II = MECHANICAL SENSORS DISPL I, DISPL H = DISPLACEMENT SENSORS ick and thin fibers. The specialized lateral line organs (electric scnsory org,-iiis) be subdivided into two main (rroups: ampullary organs and tuberous organs. :e arc many subdivisions of t]-iesc or-ans and we have previously reported about different lateral line organ types, canal organs, free neuromasts, ampullai-Y ns and tuberous or.-aiis, have a characteristic distribution pattern being similar ,-y=otids, except for Electiopl-lorus electi-ictis. The density of each type of organ 3 same species depends upon the size of the specimen, e.g., canal organs in the re 'on are separated by 1 mm in a Sternarchus -tlbif-rons 13 cm long, and by 2 mm e 22 cm long. ippro.%dmate disttibution of sing0 e organs was established by. observing the "pore r3Y' on the surface of the skin with a dissecting microscope. It follows from the ous descriptions of these organs tl-Ltt these "pores" should not be considered as I hole in the integument in every case, but rather a local differentiation of the rmis overl3,ina, the sensory or-ans V. RESULTS AND CONCLUSIONS )e was to develop the necessary instrumentation for microolectrode recordings @cti-oreceptors of electric fish so as to be able to in,%,esti.-ate their pattern ion ability. In the prenious chapter we demonstrated that the system is worl@c- and we succeeded in recording to ous autorliythmic electrical activ2ity ., the au nom nic anipull.-try clectroreceptors of Stern,-Irchtis all.)ifrons compared with the orgali's normal activity (Figures 65 and GG). rdiii- , was made during restin- of the fish with no stimulus. The impulses -egular around a repetition rate of between 100 and 300. The amplitude was 2 5 mv per spil.;e. The spike duration was very short -.around 200 micro- (r, igtire:; 63, 66). .r investimation, it can be concluded that electric fishes could use their elec- ins (trausn-iittin(r and receiniiig) for navigation ,tnd communication - in other 0 )attelm recognition. )r&,ii,-,,s and Iiistolo-ic,-LI evidence show that Sternarcl-itis -tlbifrons has three electioreceptors: an-ipull,,Lry tonic nonsy,-,ichronous units, ainpullai-Y tonic IOUs Units, and tuberous phasic nolis3rrichronous units. The physical analo-s -ind phasic electroreceptors ,ire shown i2n Figures 57 and 58. Both are repre- a generator coi-mected to resist,,tnces and capacitances in series and in The (iifference I)ctiveen tonic and phasic electroreceptors is that the first a one resist.,tnce in series with the renerator whereas the pll,-Isic electro- 9 s ha-%,e a capacitance. The tonic clectroreceptors seem to be predominant, @ze fivc-to-one, conip,,tre(I to the phasic clectroreceptors. The elcctrore- seem to act, to ,t cert,-dn extent, independently of the m.-tin electric tr-.insiiiitting least tN%,o out of three different t3-ly-,s of clectrorcccp7tors -Lre as)-licliroiiotlq -i6 .1. Figtirc 65. Recoi-clin(v from an iostlictizcd, curarizccl Stern.,trclitis imcil. Ilorizontal: 2 all)ifi-oiis si)cc ertical: 1 gr,-iduation I msec. V 1 gr-.iduatio 10 mv. n -.rv. i,-Ure 66. T@licroelcctrode recorclii-iC,) of ie autorhythi-.-iic electric. f -tl activity o ic anipullary, toiiic electroreceptors of tei-n,-irclitis -ill)ifrolis. The spil-@es seeli ali-i-io2s t n the top of the rhytliiiiic iiiusoicial %vavefori-n ire the electric icr als froiii the clectrorcceptors. n orizontal: 1 -r.-idtiatioii = 2 iiisce. ertical: 1 yriclti.,itioii 500 iiiv. .mplificatioii xl.00, offecti-%,c 1 ,-i-a(iua- ;@on 5 mV. Spike app. 2 to 5 iliv. 6G ,ae type of electroreceptor will synchronize with Ilic main electric organ. ound that the complete denervation of the transniittin.- elect-i-ic'organ does e activity of the asynchronous electroreceptors (both plilsic and tonic). The I capable of respndin- to conductive and nonconductin,,e objects placed near )ody. It may affect the total capabi2lity in determining certain movements or a certain extent, its sensitivity in pattern recocrwtion. Some of the syn- )nic units are connected to one and the same nerve trunk part of the acoustico- !stem but connected to specialized big nuclei in the brain. trildig fact about fresh water weak electric fish, besides their spontaneous in 2 ,an, is that all of them are provided with a highly developed lateralis I e slated to this acoustico-lateralis system is an enlargement of the cere- )ecially in G s niloticus and in mormyrids. The unusual importance -ilis system in tl-iese fish, compared with other teleosts, is not due to an in- aber of "ordinaiy' lateral line sensory organs, but rather to the exist2ence umber of specialized sensory organs within this same system. )ortiiic,r our hypothesis about a hybrid complex undeinvater pattern recognition I by electric fishes in recognition of prey, predators, and navigation in is recommended that the other lateralis line systems from different fresh electric fishes should be studied with the aim to find2 out the role of the isory organs in pattern recognition. of the electric fish pattern recorniition system would malce it possible to models of the physical arltlo-s of the sensors could be integrated in object 0 location, detection ,Lnd identification. The range and sensitivity of the 1 be issessed,,ind impro-,,cii-icnts could be made. lely used anaestlictic,- "INIS 22211 or "Fiiiquel" (triciiiie inctlianesulfon.-tic) - @cts the repetition rate of the electric impulses of the electric or,-,iii. It ;.o do a series of c.%-perli-nents on (lifferent aiiiestlictics to eslil)lisli AX,Ilctlier -,@e is one which tvould not affect the frequency of the impulses. It has been found thiopent,,d sodium (sodium pentliotal) N%-LII not affect the frequency of the impulses is a safe ,tnaestlictic for fish, actin(r fast and without any ill effects. -@-r subdivisions exist between the one and the same type of electroreceptor, but this 2 not been -is yet innrestigat@ed in a detailed way. The connections bet%@een the electro- aptors, the different nerves, nerve-trtmks and the brain have to be investigated. In way, their iiiterrel.-itionship could be established. Microolectrode recordings from electroreceivqrs proper and from their nerve fibers have been planned. Electro- ;ect2ion and clearing of the lateral line near the electr'oreceptors to be investigated enable us to record from the efferent nerve fibers. m the e-%-periments with Gynu-iarchus nilotictis . we concluded -.hat one fish would ognize .tnd communicate with another one of the same species. Behavioral experl- itsinthisdirectionwillbecoiitinued. Itisrecommendedthatfurtherii-dc2ro- ocle recordin-S be made from the clecti-oreceptors and from the nerve fibers of .narchtis -tlbifrons and of the newly received G@i,-i-iin,'U-clitis niloticus. 3 possible to simulate an equivalent sensory system respondin- to different stimuli m er-%vater. A system with a dou2ble feedback mechanism can be envisaged: (1) one resented by a constant frequency electric field tralismittin- system operating on phase-synchronous electroreceptors respondin- to discontinuities in the electric d or to changes in the phase relationship transmitter-r'ecel)tor; and (2) another represented by a variable freqliency tr-,insmitti2ii,- to disturbances system responding 'ie field bet-%%?een transmi e i itcr and receptor with a cl=i- of the frequency of the ismittliig electric organ. To tllis we could add 'Lii independent dual au*.Orh3,Uil-ilic 2 @-ptor system: (a) respoiiding with the increase or decrease of the autorh@,tlimic luciicy dcl)ciidin- on movcniciit directioii of the (listurb.-Ulce in the electric field; (b) responding with a cl,,an-c in ilic latency (lopcndiii- on the magnitude of the dis- ),.ince, -6dso dist-higuiswn- bet-%@,een condticti-%,c and noncondtictive objects. odel for mn,,irchtis rdloticus, one of the most sensitive electric sh s, served as a m simulation of a simple pattern rccogidtion system (Figures 67 and 68). t country of.origin: Sudan. One of the Figure .67. htis niloticus. most Sensitive tiectric Fish. (Half-scale) Sliin.41 Cot,d Lk Air 131addei, Electric ti,atl.,3mittilig oi,gans Electric receptors 3 kinds + + +* t +.+ i i i9 Electric receptors 3 kinds Electric traiisinitting organs (Electric Fish). Placement of Electric Figure 68. 232nn@@@otic@us -kransmitting and Receiving Organs. _T 0 49 AJ$4 Fig. 69 Laboratory for investicration of electric fisfies A. 7'1 1.4 4 Figitre 70 1,,iL)or;itoz-.%, f(,i- iiiv'c.;tig;itioii of clt-ctl-ie fislies 13. VI. REFERENCES N@latanabe, A., and K. Tal@:eda, J - E,,-n. 73iol.- 40:57 (1963). Bennett, 1%1. V., -@lodes of Operation of Electric Or-ans. Annals of the hTLei@? t:l York Aetdemy of Science 94:2:458 (19Gl). 2Eccles, J. C., P. Fatt, and K. Koketsu, J. Phirsiol. 126:524 (1954). Rose, J. E., and V. B. l@IoLmtcastle, Bull. Jolin Hol)],dns Hosp-, 94:238 (1954). Grundfest' I-I., Pro-7ress in Blo _1-)Tl-,,sics 7:1-85, Percramon Press, London, G. B. (1957). Furul@a%2\,a, J., and Furshpan, E. J., T,,vo inhibitory mechal-isn-is in the I%Iautlmer neurons of -,;oldfish, J. I\Teuroph3,siol. 26:140-176 (1963). Dian-ioi-id, J., and Yasarffil, G. M., S3,iiaptic function in the fish spinal cord, Dendritic integration. Pro-. Brain Res. 31:201-209 Auerbach, A. A. and Bennett, M.2 F. L., Chemically mediated transmission at a giant fil:)er synapse in the central nervous s3,stem of a vertebrate, J. Gen. Physiol. 53:183-210 (1969). Auerbach, A. A. and Bennett, 11. V. L., A rectifyiii- synapse in the central nervous system of a vertebrate. J. Gen. Pli@,siol. 53:211-237 (1969)2. A-alides, E. and Sparl:s, S., Tlio compound eye of artliro-i)ods in "Research 0 and D--velopment!' GDPD General L@-m-iiides, pl). 317-.')22 (1968). Hartli2-ic, 11. K., Wa,,mier, II. G., -,iiid l@L-Ltcliff,. 1., hi]-Libitioii in t2he eye of Liiiitiliis. J. C,cn. Pli-vc3iol. 39:G51-G7'0 (195G). Lissiiim, 11. W., ,uid T@lulliii-or, A - i%l. , Or,,,,ti-dz,,tUoii of tiiil)tlll'IIT electric receptors in G-%,iiiiioti(lic (Pisces), Pr6oc. Ro-\?. Soc. Sz,il.)o, T., S(-,nse or-,-uis of the Iitera Iiiie sysiciii iiiso)i)c electric if the j'\Toi-I)I)ol. 117:229-250 (II)C.5). 6. Szamier, R. B. and NV,-tclitel, A. W., Special cutaneous' receptor organs of fish. VI. The tuberous- and amlrillary organs of H3rpopomus. J. Miristi-tict, Res. 30:450-471 (1970). -7. De Oliveria Castro, G., Morpliolo-ical d,-ita on the brain of Electropli-)rus electricus (L) in DiO2clect"02"esis (C- Cha-as and A. P. de Carvallio, eds.), pp. 171-182, Elsevier, Amsterdam (1961). L8. Bennett, M. V. L., Meclianismof electroreception. In "Lateral Line De- tectors" (P. Calui, ed.), pp. 313-393, Indiana Univ. Press, Bloomington, Indiana (1967). 19, Suga, N.', Electrosensitivity of canal and2 free neuromast organs in a gynmotid electric fish. J. ComT). Ne@irol. 131:453-458 (1'967). 20. Bennett, M. V. L., Similarities 1),--tNeen chemically and electrically mediated transmission. Iu "Physiological and Biol)liysical Aspects of Nervous Inte- gration-" (F.D. Carlson, ed.) pp. 78-128, Prentice H-all, Englewood Cliffs, 2New Jersey (1968). 1. Mullin.- ,er, A . M., The organization of ampullary sense organs in the electric fish Gymnarchus niloticus, Tissue Cell 1:31-52 (1969). 22. Lissman, H. W., and K. E. Macbin, J. E.%cp. Biol. 35:451-486 (1958). 23. Bullock, T. H. Personal communication (1969). 24. Szabo, T. and Fcssard, A. E., J. Ph2y5iol., Paris. 57:343-360(1965). 25. Btillocl@, T. H. and Cliichibu, S., Proc. National Acad. Sci. USA. 54:422- 429 (1.965). 26. Acralides E. Evolution of electric or-ans in fish, information processing by 0 0 electric fish and communication beinveen electric fish. Lecture at the Arthur 2 D. Little Co., Caml.)rid(r ,C, I\Iass. (21 Feb. 1961). 27. Arrlilides, E., D. Hcl)l)ncr ,t,.id C. Claysi-iiith. Communication techniques to 0 resist jamming. GDCA-ERR-1717, Coii-%rair Aerospace Div. (Octol)cr 1972). 28. Hul)cl, D. H., Scic0iiccs, Vol. 125, pp. 549-550 (1957). 29. NN'ollib,,trsht, M. L., l@,lacN!chol, E. F., Jr., aiidW-.tgner, 11. G., Science, Vol. 132, pl). 1309-1310 (1960). ry, R. W.# -irundfest, II., Scngst.,.Izcn, R. W., Oettinger, W. II., and Giir IC,. of -,gci. Tnst-r., Vol. 21, pp. 3GO-3Gl (1950). -)on-,adson, P. E. K., Electronic Apparatus for Biological Research, Butter- vorths Sci. Publ., London, pp. 568-581 (1958). Vood, E. 1%1. , Urethane as a carcinocr n. Progressive r, ish-Culturist, Vol. 18, C)C 4o. 3 (1956). 3all, J. N., and Cowen, P. N., Urethane as a carcinogen and as an anaes- hetic for fislies. Nature, Vol. 184, p. 370 (1959). 3ove, F. J. IIS-222 Sandoz, the anacsthetic of choice for fish -Lnd other cold- )Iooded organisms. Sandoz News No. 3 (1965). ,Iarldng, L. L. , Investi-ations in Fish Control: 12. Toxi. ity of AIS-222 to t), ;elected fislies. U.S. Bureau of Sport Fisheries and Wildlife Resource Pub- ication No. 18 (1966). @Iall@er, C. R., and Sclioett-er, R. A., Investigations in Fish Control: Resi- t@ iues of MS2-222 in four species of salmoiiids following anesthesia. U.S. Bitreati ildlife Resource I>til)lication No. 21 (1966). '@choett,a,,er, R. A., and Julin, A. M., 13. Efficacy of MS-222 as an Anesthetic ,n Four Salmonids. U.S. Bureau of ort Fisheries and. Wil life Rgsotirc_e ')ul)lication No. 19 (1967). :lont2z, G. W., "Anesthesia of Fislies" , in Experimental Animal Anesthesiology )y D. C. SaNvyer,, AD 648 329, U.S. Dept. of Commerce 19G7 Symposium held 3ec. 1964 at the Brooks AFB, Texas. .he I%IERCK INDEX. zabo, Th., in Lateral Line Detectors (P. Calin, ed.), pp. 295-311, Indiana iniversity Press (1971). lagiwa.ra', S., Szabo, Th., and Engor, P. S., J. NcuK2phyEi2L 28:775-7S3 (19G5). '.ncrer, P.S. , and Szabo, Th., J. of Netiropli,%,siol. 28:775-783 (19G5) ,obu2L Suga.,-J. of Conip. Neurol. 131:4:437-451 (1967). 'ulloclct tin Bch,.tv. Evol. 2:85-118 (1969) T. 11. Br, lartop: U. S. Patent 3109001 to Abbot Labs rlg6@). @galides, E., Sensiti,@ity and Behavioral Reaption of Sharks to Electric ')timuli, Final Report. ONTR Contract Nonr 4773(00) No. 104-8G3 (1967). lagiavara, S., Szabo, T., and En,-or, P.S., Electroreceptor mechanism in -t high f@requency wealdy electric fish, Stern,-t6rclitis albifrons. J. Nourophysiol. ),'8:5-.784-799 (1965). Szabo, T., Nature, 194:4848:600-001 (1962). 7 ri