THt SENSITIVITY AND RESPONSE OF WEAKLY ELECTRIC FISH TO STATIC AND PULSED MAGLIMTIC FIELDS June 1, 1970 I. ItiTRODUCTION. The study of bioelectrocenesis, particularly --n the various species of electric fish, has been of increasing scientific concern in recent years. This interest stems primarily from the potential usefulness of research in this area in contributing to our understandina of a number of fundamental and significant problems. By defining the e2lectric fish's unique sensitiv- ity to electric and magnetic fields, and how it codes and utilizes such sens- ory information in its detection and navigation behavior, current evidence is providing a more complete concept of such basic'questions as migration and territoriality, and is leading toward the deve'Lopment of various bionic devices in the form of underwater sensors and power sources. In ad2dition, knowledge of the effects of magnetic and electric fields on physiological and behavioral processes has assumed great importance in view of man's ex- posure to drastic changes in such stimuli during space travel. Living things produce a changingelectric field at and near the surface of their bodies; all fish, being sheathed in a conductive substance and living in a conduct2ive medium, produce an electric field that may be detect- ed at relatively great distances. However, there are certain fi'sh which produce electric fields exceeding the norm by hundreds or thousahds of degrees of magnitude. The electric eels of the Amazon can 'produce bursts in excess of 600 volts. Other electric fish, i.e., weakly electric fish, produce continuous fields measured only in millivolts, but by 2means of in- terpreting distortions in these fields are able to sense and navigate through their environment to a degree comparable to that of other species in which vision '@s used for these purposes. The weakly electric fish, having very poorly developed visual abilities, must depend on i,-Iformation acquired through their electric fields in order to survive. The mag-ie2tic field is a form of energy to which all plants and ani- mals@are exposed., Its influence on living systems, however, is subtle and not well understood. one approach to studying the effects of magnetic fields upon behavior is through the use of an organism which produces an electric field and uses it as a detection and navigation mechanism. The electric fish is just such an organism, and this report2 will review a one- year study of two of these species (Sternarchus albifrons and S. ler)tor- hyachus; see Figure 1) which has just been completed In these fish, impulses are dis- charged from the tail and received by the head, which becomes positive in regard to the tail. This potential difference creates an electric field 2 about the fish's body, permitting it to detect objects through distortions in the field. Several studies have shown that these fish can perceive a static (constant strength) magnetic field, but only when either the orga- nism or the field is in motion, thus generating a current in the fish. It was thought that the fish was responding to the current generated in itself by the magnet. However, in these expe7riments the magnetic field was presented as a static field, and the sensitivity of the fish to a pulsed field presented at various frequencies, particularly the frequency at which the fish discharges its own electric field (500-1500 cps), was not investigated. Other investigators have shown dramatic increase in sensi- tivity to Ap2lied A.@g. approximating the frequency of the fish's discharge. In addition, the strength of the field was not systematically varied in terms of the gauss level in the fish's proximity. Therefore,2 there are considerable gaps in our knowledge of the degree of sensitivity of the fish to magnetic fields at various frequencies and strengths. The present stua-,v was undertaken to clarify sou,.e of these problems regarding the perception of and response to a magnetic field which is systematically varied along several continua. Figure 1. Ster-iarchus albifrons (t2op) and Ster-narchu evtorhvnchus (bottom) !us I IT. BACXGROLND. Comparatively little work hag been done on the sensitivity of weakly electric fish to various types of electrical and magnetic fields, although the evidence that is available indicates that these fish have an extremely low threshold for such stimuli. Liss=nn2(1958) and Lissmann and Macnin (1958), for example, have shown that gvmnarchus niloticus will perce've the movement of a magnet or an electrified insulator when either is Moved outside its tank or aquarium.* A small bar magnet was held against the *Szabo et al. (1969) w-rite that the electzqreptors respond to both the presence and movement of an object or field. 3 2 wall of the aquarium and mved in avertical direction, with the result that a "single downward sweep produced aresponse in the fish if the rrovemnt was sufficiently rapid and the distance between the fish and the magnet suff-i- ciently small. With the particular magnet used a response could be elicited at a velocity of about 3 m/sec when t2he fish was about 50 cm from the magnet" (Lissmann and ifachin, p.451). When an electrostatic charge* was moved hori- zontally in front of the tank, the fish was seen to respond to a voltage of 60 kV when the distance from the fish was 50 cm and the charge was moved at 3 m/sec. The authors conclude that Gvmnarchus is able to detect potential gradients of about 20.30/A@/cm in the surrounding water. Table 1 shows the remarkable sensitivity of this species as compared to other fish. It is apparent that the perceptive ability of Gymnarchus is of a different order Table 1. The sensitivity of six species of fish to direct current. (After Lissmann and Machin, 1958.) Current density 2 Species (,uA./cm.2) Phoxinus 2h2Linus (minnow) 10 Cypriniis cardio (carp) 60 C. auratus (goldfish) 16 -ia@a-siiu-rus, asotus (catfish) 2 8 - Ca@;-terosteus acut-eatus (stickleback) 110 2ymnarchus niloticus 2 x 10-5 of magnitude than other fish. Since it can detect a direct current of about .15 microvolt per centimeter, an individual sense organ in Gymnarchus should be sensitive to a current 2change as small as .003 micromicroampere. Lissmann (1958) has observed that Gvmcrtus carat)o can be conditioned to feed in response to a stationary permanent magnet mounted outside its tank and to inhibit feeding responses when the magnet is absent. He notes that although there is no sipecifically relevant data, it would seem that this fish should be able to2 perceive a field of about 10 oersted when moving at a rate of 10 cm/sec. In a subsequent paper by Machin and Lissmann (1960), it was shown that the receptors responding to small direct currents were also used in the fish's object detection and location. That is, "the sensitivity of the fish to ex- ternally applied currents gives information about the electric recept2ors for object location 11 (Machin and Lissmann, p.802). *A small aluminum cylinder on an insulated handle and charged from a Wimshurst machine was used. 3 A. liagnetic field effects. The effects of various types of magnetic -.@ields on living organisms has been a subject of increasing interest in recent years for both theo- retical and practical reasons. "Basically, the magnetic field, being a form of energy, just as are light, heat and sound, impinges upon all liv- ing or-anisms whether plant-2or animal. The question as to its effect on living matter is what we are seeking to learn. Is it an active or passive process? How will an organism react tD an environment that is devoid of a magnetic field? Further, what will happen if the field is altered or distorted?" (Caldwell and Russo, 1968, p.233). Caldwell and Russo studied the effects of an A.C. magnetic2 field upon the behavior of the Italian honeybee (Lois mellifica), and found that the bee would respond to the magnetic energy field with a stereo- typed nodal reaction, i.e., three of the four subjects would situate themselves and become riaidly fl-ated over one of the magnetic nodes When the magnet was on. Cottlieb and Caldwell (1967) investigated the ma2gnetic field effects on the compass mechanism and activity level of the snail Helisoma deyi endiscui. Using a bar magnet with a weak field (1.5 gauss), they obtained significant effects on the activity level of the subjects. Since astronauts have and will continue to be exposed to magnetic fields -.4hich are much less intense than the Earth's magn2etic field while exploring the surfaces of neighboring celestial bodies, "the question arises as to whether the human body has during its evolution become de- pendent on the presence of the Earth's magnet-r@c field for the maintenan,:e of its normal functional integrity. Accordingly, it has become most im- portant to ascertain whether a low-intensity maanetic field exposure 2 could pospibly lead to an impairment of health or performance of an in- dividual" (Busby, 1967, p.7). However, there is also the possibility that astronauts Could be exposed to intermittently high-intensit-/ mag- netic fields up to 1,000 gauss for varying periods during space travel. Beischer (1963, 1969) and Beischer et al. (19672) have studied the ef- fects of both low- and high-intensity fields on man and animals. Their results show that man does not seem to be affected by a two-week exposure to 50-gama fields; mice survive a one-hour exposure to 120,000 gauss; and in a low-intensity magnetic field, there is a significant gradual decrease of the scotopic flicker fusion thres,iold in man. 2 Agalides has recently completed a series of studies on weakly electric fish, including some work on their sensitivity to moving magnetic fields. Using GXmnarchus and Sternarchus as subjects, he observed that they responded to a permanent bar maonet of 930 gauss. The magnet was moved at 3 m/sec and was perceived by the fish at a distance of 120 cm. This was 0very close to the fish's sensitivity to static electric fields, and corresponds to a gradient of 3 emu, or 0.03,tAV/cm. 4 B. Electrosensitivity Granath et al. (1967) worked with Sternarchus albifrons in their ef- fort to determine its sensitivity to imposed electric fields. To study the frequency response continuum, the authors used a conditioning prob- lem in whizh both uniform and nonuniform alternating current (A.C.) fields were employed as signals for the subjects 2to leave a porous cylinder and swim to a vertical plastic tube for a food reward. After the conditioned response was established with high stimulus values, the signal was re- duced to daternine the threshold of the fish. T@,e results indicated that Sternarchus is most iensitive at its own discharge frequency at room temp- erature, i.e., in the area of 1,000 cps, wit2h a maximum sensitivity of 0.2 microvolts per cm. However, a secondary maximum was observed at the second harmonic of the discharge frequency. Watanabe and Takeda (1963) employed the South American gy=atid, Eigenmani , in their study of the effects of ekternally applied electric current. Like Granath et al., they found that the effective stimulus was an2 alternating current presented at a frequency very close to that of the fish's own discharge. In this case, Bigennania has a discharge rate of about 300 cps at 250C. Their results showed that "when a sinusoidal (or a square pulse) electric signal with a frequency similar to that of the fish's own discharge is applied to the fish, the latter's discharge frequency changes aG if to escape fro2m the applied signal frequency. The effectiveness of the stimulus depends on the difference beween the two frequencies (.!@S); whenZ@S is more thanlo cps the response is barely rec- ognizable. The smaller LS, the more effective the stimulus, except when AS is very small, where the response again fails to occur" (Watanabe and Takeda, p.65) Dewsbu2ry, (1966b) believes that stimuli* differ and interact in the kind and/or amount of change they induce in the discharge frequency of weakly electric fish. Hie observed several different species, but not Sternarchus albifrons which does not appear to behave in this way. Dewsbury attempts to relate his data to a concept of arousal, wherein discharge frequency changes with arousal level. In anot2her study (1966a), he confirmed the hypothesis that electric organ discharge frequency in gymotids is higher in darkness than in liglit. This would normally be expected, although we have not found such evidence in S. albifrons. The effect of temperature on discharge frequency is a particularly important problem in that'the exact nature of this relationship must be 2 knl-wn in order to establish baseline data for further study on the fish's discharge behavior. Gallon et al. (1967) and Enger and Szabo (1968) have found that the rate of discharge varies with temperature in mor:nyrids and *For example, light-darkness, shock, aeration, metallic objects, and a buzzer (Dewsbury, 1966c). 55 gymnotids, Their results are summarized in Figures 2 and 3. I 0- 030/20 2 2 03 02 Lli 030120 2..i- 2 0 12- G3 030/20 a? 4 6 1 6- 0 t 2 20 25 30 TEMP. "C Figure 2. Discharge rate as a function of temperature. Open circles, as- cending series; filled circles, return to lower temperature; triangle, second ascending measureuznt. (After Gallon et al., 1967.2%, 2000 Gioll L3 5m a 06 Sternapylus 2 E Increasing te". Decreasing terr4,)@ 20 25 30 35 IC Fisure 3. Relation between water temperature and di1scharge rate. Open circles, increasing temperatures; closed circles, decreasing temperatures; broken lines, Qlo- values for comparison. (After Enger and Szabo. 196a.) Attem@)ts have also been made to condition the discharge rate of morntyrids with both classical and operant methods. Mandriota et al. (1965) report that three species of Mormvridie would briefly increase their dis- charge frequency (conditioned response) in response to light (conditioned stimulus) following training trials in which light was p2aired with shock (unconditioned stin-,ulus). Mandriota et al. (1966) later discovered that operant (avoidance) conditioning was also effective in these fish and, in fact, was uiore efficient than classical conditioning in that fewer shocks were required to establish the response. 111. CUR-RENT STUDY: THE SENSITIVNY AND RESPONSE OF STERNARCEUS ALBIFRO'tqS TO STATIC AND PULSED 14AGIqETIC FIELDS, A. Problems and hypotheses. Tl,.e prixaary hypotheses of this study were concerned with the problem of determining whether weakly electric fish are sensitive to magnetic fields and, if so, how this sensitivity might vary as the field is changed from a static to an alternating and to 2a pulsed one, as the frequency of the field is increased or decreased in relation to the normal discharge frequency of the subject, and as the strength of the field is varied., A secondary problem conceined with effects of various drugs on the elertrical activity of the fish was also investigated. H(xiever, before the data could be collected, it was necessary to find a 2source from which weakly electric fish could be obtained, develop life-support systems for the subjects, and design and construct the required equipuent and apparatus. le Subjects. A total of 18 fish were purchased, consisting of 9 Sternarchus albifrons, 6 S. le2torhvnchus, and 3 weakly electric fish of an un- known species. Most of t2he specimens were bought from the Cappet Corporation in Alexandria, Virginia, and a few from local pet shops. Four of the fish, all S. altbjifr., remained healthy during the peri- I.L.trons od of the study, and were the only ones used in the final experiments. Viair size Is shown in Table 2. Wit2h few exceptions,.the others died within one week of purchase, Table 2, Size of the four experimental S. albifrons. $,4bjict Length lish #1 18 cm Fish #2 21 cm Fish #3 14 cm 7 Fish #4 11 cm 7 The subjects were kept in individual tanks of either 14 or 20 liter capacity. The water was aerated with conventional air pumps working through ctiarcoal and glass wool filters. I'he temperature was maintained at about 26.SOC and the pH at 6.7 to 6.9. Food con- sisted of either live or dehydrated brine shrimp. The fish were fed 2 2 - 3 titres a day, and once a week an antibacterial agent was added to the water to suppress the growth of bacteria. 2. Equipment and apparatus. A plastic Y maze (Figure 4) was constructed for tests with the the static or steady ma@netic field. Its three arms were joined at angles of-1200, and the maze itself made of 0.04011 sheet styrene 2 fastened with styrene solvent. The water in the maze was drawn from a continuously aerated and filtered source with a pH of 6.8 and a temperature of 26.80C. It was exchanged every @ hour, within which tirrie the temperature drop was approximately 0.20C. However, because of its inadequate size, this maze proved unsatisfactory, i.e., the larger subjects co2uld not be used in it. In addition, with a magnet- ic field of 8-10 gauss and the magnet centers at the distal end of one test arm, a minimum field of 2 gauss was present at the farthest point in the starting chamber. The field was 4 gauss in the area of the fish's head when the subject was released from the starting cham- ber. These problems with the Y maze may have contributed 2to the failure to find any response by the fish to the magnetic field in the initial experiments. A larger T maze (Figure 5) was then designed and constructed in an attempt to demonstrate a more positive response to one arm at the choice p@,.int with the magnetic field as a cue. In the Y maze, the subject appeared to swim into the arm on the side of the 2starting chamber that the fish was closest to when the door to the starting area was opened. The length and depth of the T-maze arms were much greater than those of the Y maze, permitting the use of the largest specimens. This maze was also provided with continuous filtration and heating. It had a 10-liter capacity,'with an auxiliary 16-liter and 50-2watt heater. An air lift and siphon were arranged so that heated, filtered water was slowly and continuously fed into the leg of the T from the tank. A return siphon ran from the distal end of each arm to the auxiliary tank. Temperature in the maze was held at 26.8'C by maintaining the temperature in the external tank at 32.50C. The tank and siphons were wrapped with paper 2lagging and jacketed with aluminum foil to achieve and hold the desired temperature. The maze itself was made of kll sheet clear acrylic plastic, joined with an appropriate solvent. The material proved to be light and strong, and permitted good observation of the fish. The magnets were made by winding 5 pounds of #12 copper magnet wire on each of 2 alu8minum tins of 23 cm diameter. When in use, the 8 POWER SUPPLY 6-12 volt 20-10 Arnp. battery eliminator Y-MAZE Constructed of.040" sheet styrene Water In-the maze drawn and replenished half-hourly from 80-degree F-7. 9 piff source. Temp2erature loss la the Interval negligible. STIMULATOR 9 volt 22 mA@n thru stialnless steel electrodes Figure 4. The plastic Y maze shown with the equiprent (tuagnets. power supply and stimulator) used in tests 6 of the static magnetic field. v eE o n= version of "rs d vo] cm:indtjc:e ive @@7.5cm@@-15cmE,8 ced wave 30cm 45cm @c m 4L ma net 10cm 0.24V 0.2V 8OmV -12mV -5mV 24G 2OG 8G -1.2G -5G 2 Distribution of 14agnetic Field' Voltages shown are those i@,luced 41 and form of induced puise in in a 300 turn test coil of 3cm dii tb,e Electric Fish Tee Maze.- 25cm 2 @2@Smvp2e5'G 16mV, 1.6G 5 cm. (110 Io 5niV,0.5G 7@5@ dep a__ cm) 1 6=7 th 3. 'Ahe mnd the of -t@%,z @4"lene, '-,n It. 2 coils were wired in series and placed on either side of the maze arm, 13 cm apart. The discharge patterns of the fish were recorded by means of probes of pure carbon rod. Above the water level, these probes were shielded in thickwall aluminum tubing. Shielded cable was used to connect the probes with the oscilloscope input. The oscilloscope 2 was a TEKTRONIX 502 A. and the built-in prcamp was found to be suf- ficient to record these fish at distances over 50 cm, which exceeded oixr needs for these experiments. In recording, the probe shields, the cable shields, and the scope ground were all connected to a doub- le wrap of heavy aluminum foil arounl the chamber containing the fish. With this 2arrangement the "noise" level on the system was held to 0.3 mV, which was-acceptable. The electrical equipment which surrounded the experimental ap- paratus was a source of an electrical noise electromotive force (E'MF) wiich drove current through the input resistor R I of the IP input$ 2 measuring device (in this case an oscilloscope).. The noise power dissipated in such an input resistor is a constant, Pn, so the noise voltage developed at the input of the scope is Vn=FPnRi.,ut (1) and therefore by reducing the input impedance "Rinputo noise voltage is reduced. The network shown in Figure 6 reduc2ed the input impedance of the oscilloscope from its usual value of 1 @leoohm to 20 kilo-ohm 0 and allowed the input imped3nces for the two separate beams to be balanced in order to eliminate any assymmetries in the external net- work. The noise improvement achieved by this method is a factor of 2 7; the measuremnts were not affected since the impedance of the source electrodes and tank was about 500 ohms. As long as the source impedance is low with res@ect to the input impedance of the masuring device che source is not affected by the measurements. The operation of the device is straightforward. After all of the shields have been conn2ected, the 40K potentiometer is adjusted until the noise on both beams of the oscilloscope is minimized. This is the bes't operating point. The distribution and strength of the magnetic field were determ- Lned with a test coil of 300 turns, 15 mm in diameter. 'The induced electromotive force.(EMF) was converted into a field strength measure- 3 ment in gauss by application of Faraday's law EHF (2) dt ,where is the magnetic flux through the circuit. Equation (2) says that the E@IF induced in a circuit is equal to the rate of change of the riagnetic flux through the circuit. The magnetic flux is Beam #1 input. to electrode 1 m 40 K Oscilloscope to electrode #2 I m shiela Beam #2 input 6 Figure 6. Input impedence'reduction network. 12 as <> been added. By varying the voltage output of the power supply, the voltage output at the si-nal generator, and by careful adjustment of 0 a variab:e bias-resistor at the simnal input to the transistor "switch", 0 a pulsed field of the desired characteristics was achieved. The2 circuit schematic is shown in Figure 7. The circuit consists of four npn transistors, three of which ('/'2N3055) switch all of the current through the magnet, and one (#ZN3054) receives the signal from the signal generator (Lafayette #99-5014) and drives the three power transistors. The resistors in the circuit are bias resistors and the capa2citors tend to round off the switchin- ptilses and prevent oscilla- tions. When'the sinxisoidal signal from the signal generator goes posi- tive the transistor switch turns on and the current flows from the 12V poi;er supply through the magnet. When the signal reaches the negative portion of its cycle, the switch turns off and current is 2 prevented from flowing. By putting in a 1000 Hz signal we therefore put 1000 pulses per second through the magnet with a maximum of 12 amps peak current. The actual current through the magnet is considerably less since the inductance increases the im-pedarce. The transistors dissipate a great deal of power and must be placed on heat sinks in order to oper2ata properly and to prevent thermal failure. B. Experiment 1: Temperature-frequency baseline data. We had observed that the discharge frequency of each fish at a given temperature was different from that of the other fish. Agalides reports that temperature-related frequency changes in 'S. albifrons-are complex, but are on the order of A50 cps/&CO, which our work confirme2d. Since the current experiuents required an accurate prediction of discharge frequency, a study was made of the temperature-frequency relationship in the fish in order to provide baseline data. Each fish was monitored for frequency at '10-12 points in the 21 to 310C range and over a period'of 6 weeks. The test chamber was a plastic box 25 x 43 x 20 cm, double wrappe2d on the exterior with heavy aluminum foil. Three liters of water were drawn from the test fish's home tank and placed in the box. Temperature variations were achieved with a heat exchanger made from a plastic pitcher and a length of plastic tubing. The tubing was coiled in the pitcher, which was filled with either hot or cold water. Water from the test apparatus was forced through the 2 tubing at such a rate as to change its temperature 1OC/15 min. When the desired change was achieved, as determined by an electric thermotreter, 2 minutes were allowed to elapse, and then the frequency of the test fish was recorded. Before recording, the sensing thermistor of the thermometer was removed from the apparatus because it introduced extraneous signals into the water and, thus, i8nto the oscilloscope used for frequency de- terminations. Fifteen-minute observation periods indicated that frequen- cy always stabilized in less than 2 minutes. 14 Magnet +12 v 150K 3055 @Ilt4 055 3i9gnal input 3054 055 *Note: The'2N3055's may be replaced by the more costly, SK3027 Figure 7. Schewtic of magnet switching circuit. 15 A series of observations with the fish maintained in close fitting rigid styrene tubes showed no variations in either che amplitude or the phase relationships of the discharge accompanyina the change in frequen- cy. These results are not in keeping with those of Agalides, who reports aviplitude changes. Within the limits of accuracy of our test situation, we fo2und these fish to have straight-line plots of temperature-frequency response with a range of +15 to +50 cps at any given point, depending on the fish. Whether thi7s varia7tion resulted from individual differences or variability in the method is not known, jut the experimenters lean toward the latter interpretation. Our laboratory was by no means 2terrperature controlled, and the fish may have been responding to changes in temperature over the- entire apparatus, which was not apparent in the small area actually sampled for temperature. The results of this experiment are shown in Figure S. C. Experiment 2: Response to unpulsed magnetic fields in the Y maze. 1. &nvironmental preference using a static magnetic field. 2 The equilateral Y maze was initially used in an attempt to dem- onstrate a sensitivity to a relatively strong static magnetic field in the sirall specimns of S; albifrons and S. leotorhvnchus. It was thought that these fish, with electric iields having a maxi potential as observed in our lab of only 6.2 m volt, and an ability to detect one an2other by mans of these fields at distances exceed- ing 1 meter, would respond.(a) to changes in this field induced by • large magnet, or (b) to currents induced in their bodies by such • magnet. However, no gross responses from the fish were observed in either swi=ing behavior or in electrical discharge pattern when the magnetic coils were arranged so that a magnet2ic field calculated at 9- 10 gauss was centered in a 40 x 20 x 30 cm aquarium iA which a fish had been previously placed. Consequently the Y maze was used- for further experimentation. The dimensions of this maze allowed relatively low levels of the magnetic field in the first 5 cm of the experimental arm with inten- sity increasing to a maximu2m of 9 - 10 gauss at 11.5.cm. The overall dimensions of the apparatus allowed minimal swirmning room for the four smallest fish: two albi,frons and two leptorhynchus 15 - 15.5 cm in length. The current induced ove-r a short distance in the environment by a standard 9-volt transistor radio battery proved to be a noxious stim- 0 ulus, and electrodes were installed at the starting point in the event that subjects did not move rapidly to the choice point. The subject was placed in the starting arm and a short period al- lowed to elapse. Both experimental species were passive fish and 16 TEMPERATURE-FREQUENCY REIATIONSHIP Z9 PLOTS FOP. FOUR STERNARCHUS ALBIFRONS Amok !8 t7 -26-.8- t6 2 s 3 t5 !4 @3 6 0 6 800 v 0 1000 FITEQUENCY Or DISCIIARGE CPS Figure 8. 'A"he disc,%zrge grequency-te,.iioerctLirp- short-accomodation periods of about one minute were sufficient before the door to the choice point was opened. The subject was allowed 2 seconds to move to the choice point. If this had not occurred at 2 seconds, the experirentor made contact in the stimulator circuit. Out of 120 trials, this was necessary only about 10% of the tim ip 2 largely with one particular fish. Stimulation once initiated was - maintained through the trial. Tne fish is not greatly affected, if at all, except in the area directly between the electrodes, but in order to control the possible effects of other variables, this tech- nique was used. The magnet was kept at the left arm, and was left on for 10 2 trials, off for 10 trials, and then on for a final 10. After a trial the fish was allowed to return to the starting chamber by the process of blocking the unoccupied choice arm, waiting till the fish had moved from the other arm, blocking it, and then blocking the start- ihg chamber as the fish returned to it during normal exploratory be- 2 havior. These fish are nocturnal and exhibit continuous searching during the dark hours. The hours preceding ard during experimental sessions were dark with only low level red illumination. The inter- val bet-deen trials was thus variable, but the fish was kept in an unexcited state. Elapsed time for 30 trials was about 30 minutes. T'L2ie results of this series of tr@.als (Table 3) indicated no sig- nificant preference or aversion for the stati,c magnetic field, al- --. though the subjects did tend to turn left in the maze. The mean per- centage of left turns with the magnet off was 52%, and with the mag- net on 54.3%. Table 3. Preference trials in the Y 2maze' with the static magnetic field. Choices to the left (Magnet on the left arm) Magnet on Magnet off Magret on Fish 10 trials 10 trials 10 trials S.A.31 30% 50% 60% 2 S.A.4 40% 50% 40% (magnet off) S L 22 50% 60% 70% S:L:3 70% 60% 60% 'nis fish required stimulation on the lst trial. 2This fish required stimulation 10 times (3,4,3 distribution) 2. Co2nditioned response to the static magnetic field., In a futther attempt to obtain some indication that S. albifrons is sensitive to a non-pulsed magnetic field, a conditioning technique was used in which the presence of the field (conditioned stimulus) was paired with electric shock (unconditioned stimulus). 18 The S was selected on the basis of size, i.e., the most suitably sized fish for the experir,.ental chamber, which consisted of one arm of the plastic water-filled Y maze. Electric shock was administered from a 9-volt battery through electrodes fastened to the walls of the chamber. The two magnets were placed on either side of the arm and activate2d by an 8-volt, 9-amp power source. When the fish was placed in the chamber, the magnetic field (9 - 10 gauss) was turned on, and k second later, the electric shock was administered for a period of one second, at which point both stim- uli were turned off. The S responded in a characteristic manner to the shock with a "startle" reaction (unconditioned response). I2t was hoped that after a sufficient number of trials the S would respond (conditioned response) in this way to the magnetic field alone, or, when both stimuli were used, would anticipate the presentation of shock by responding to the field in the initial 12-second interval. However, after four series of 25 trials each, giving a total of 100 trials, the S fail2ed to show any response to the magnetic field. Con- sequently, there was no indication that the fish was able to perceive the non-pulsed magnetic field. 3. Conditioning with the magnets at reduced intensities. S. albifrons #3 was confined to one arm of the Y maze. Two electrodes were fixed to the sides of this arm, and the magnets placed on either side. In 2contrast to the first experiment in this series, the magnets received only 4 volts and 4.8 amps from the power supply, producing a magnetic field of considerably less intensity (3@ gauss) than that used previously. In the first experizrent, the fish failed to respond to the magnetic field, i.e., it gave no evidence of sens- ing the field at full strength, and it was decide2d to attempt another test with the field at half strength on the possibility that the orig- inal was too strong, thereby interfering with the fish's afferent processes. With the fish confined to one arm of the maze, the =gn@tic field was turned on for a period of two seconds, and after the first second, e'-ectric ahock ( gvolts, 200/tfi@m2) was a2dministered to the subject for one second. At the end of two seconds, both the field and shock were turned off. As expected, the fish responded to the shock (uncondit-'-oned stimulus) with a "startle" movement (unconditioned re- sponse), but after 50 trials, when the field (conditioned stir-ulus) was used by itself, there was no anticipatory conditioned respo2nse. It appears that the fish did not sense the magnetic field as presented. A second series of 50 trials were then run with S. albifrons #4 with the magnetic field power source at 2 volts and 1.8 amps. The re- sults, however, continued to be negative. But over the course of the ttials, both fish showed sme habituation to the electric shock, which 2 had b en reduced with a recalibration of the variable resistor to 100 ,,RA/cm at 9v for the second set of trials. 19 D. Experimnt 3: Drug study. In contrast to various reports in the literature on other gyrmotid fish, no stimuli to which Sternarchus would normally be exposed were found to affect their discharc,e frequency. Such things as noise, physi- cal manipulation, light, dark, feeding, starvation and illness failed to change ,he f2reque-.icy of the fish in the current study. As we have seen, frequency changes with temperature in a highly predictable manner, and Watanabe and Takeda (1963) demonstrated a response to applied AC current. They found that AC at the fish's own frequency caused the fish to alter his own frequency in response. The greatest relative changes occurred 2 when the applied current was closest to the fish's own frequency. In an effort to determine the degree of stability and control the fish is able to maintain over its discharge frequency, a study of the effects of various drugs on their discharge patterns'was undertaken. Two depressants, Nembutal (sodium pentobarbital) and Pontocaine (tetracaine hyd2rochloride), and L-dopa (levodopa) were tested. Nembutal at 750 mg/liter anesthetized the fish with no effect on the amplitude ot frequency of their discharge. The L-dopa effects are discussed separately; they did not shoir a direct effect on frequency. Pontocaine, however, modified the fish's discharge frequency. At a con- centratio2n of 3 m- in 500 ml water, the discharge rate dropped 140 cps. Twenty minutes after a final total dose ofl.12 Sms/500 ml, the discharge was 375 cps below the expected level. The fish rested on its side and vas unresponsive to stimuli. At this time, the amplitude and phase re- lationships of the discharge were unaltered. The results are summarized in2 Table 4. Table 4. The effects of Pontocaine on discharge frequency. rugge@ Norma 'c2y Frequency Tim Dose Tem;i. _lfrequen Remarks 09:25 0.375,cm 27. 10 2 980 + "Zi:30 27.5 839 09:37 0.375gm 09:43 27.35 787 965 + 30 09:45 0.375gm 09:51 09:56 27.2 649 955 + 30 10:02 9 200 ml water re- placed with fresh, fish letl--largic 10:08 27.1 575 950 + 30 -io . i-i fish lying still In view of the considerable interest in the neurotropic drug levod'opa (L-dopa), and because the experimental techniques developed in the current study are capable of providin- direct telemetric evidence of nervous sys- tem functions, it was decided to test L-dopa on selected fish in order to determine its effects on their electrical discharge patterns and be-2 havior. The first subject was S.A.,';-4, who received the drug for 27 days. Long exposure to relatively large doses is necessary to produce behavior- al effects in other species. A technique of repeated injections as one way of achieving this was ruled out as impractical; therefore, a method for dissolving L-dop4 into the aquarium water was 2worked out. Dar7 pro- vided by the'manufacturer of the drug (Iloff,-nann-LaRoche) indicated that L-dopa is not very soluble in water; approximately 0.4% at 800F. An air- lift was arranged to bubble aquarium water from the 20 liter home tank at a very slow rate through a chamber containing the drug and lined with filter paper. The charco2al was removed from the tank's filter. Every day, weekends excluded, 100 mg of the drug on fresh filter paper was placed in the dispenser. lie found that L-dopa, under these conditions, rapidly combined with other substances present to form a heavy, dark, flocculent precipitate, which clogged the tank filter and the filter paper in the dispenser. The principal change was a conversio2n of the dopamine to melanin, which was later prevented by the addition of 50 mg of ascorbic acid every time L-dopa was added. S.A.#3 also received the ascorbic as a control. No unusual behavior or alteration-in the form or amplitude of the electrical discharge of S.A.44 were noted for 14 days. Then an increas- ing disorientation, reduced abi2lity to find food, and abnormal discharge frequencies were noted. On the 21st day of drug experimentation a tho- rough series of temperature-frequency studies was performed in the pre- viously described manner. These revealed that the slope of the temperature- frequency plot was unchanged, but whereas a variability of + 50 cps had been previously noted, varia2bility was now found to be + 150 cps as test- ed over a period of 3 consecutive days. The drug was s@opped at the end of this time, the water in the tank changed, and charcoal filtration re- sumed. Four days after-t:his, the temperature response was still quite erratic. The next close evaluation came 50 days after cessation of the drug. Temperature-freque2ncy response at this time had returned to the pre-drug parameters. The cotitrol, receiving ascorbic acid alone,-showed no such effects. S.A.#2 was given L-dopa and ascorbic acid over 47 days. The dis- penser in this case was a plastic funnel suspended with the narrow end of the cone submerged; discs of filter paper, folded in half twice and 2 opened to form a cone lined the funnel. Fifty mg each of dopa and as- corbic acid were placed into the cone and were dispersed by simple dif- fusion over a period of 2 days. This was a more reliable, less trouble- som method of dispensing the drug than that used earlier. In this case, the tank filtration was left intact, but the air flow to the bubbler 2 21 was reduced, lessening the water floi4 throu-h the charcoal. We assumed 0 that with an undissolved supply of the drug at hand, an equilibrium would be achieved, and the amount of drug in the water would be constant despite uptake by the fish and/or the charcoal. Several tests were tried, removing the fish thro2ugh 2 changes of dis- tilled water which had been aereated and brought to the proper condition with reagent grade chemicals, to test for metabolized dopamine with fer- ric chlor-'@do. However, there were no differences demonstrated in the color or quantity of precipitate betweer. experinental and control fish. S.A.#2, during and after 47 days on the drug, showed no al2tered be- bavior and no change-in his electric field. The drug studies were terminated at this point. The exact cause of S.A.#4's reactions are not definitely known. Because of his sma.11 size, the drug may have had more effect on him than on S.A.#2. He may have suffered an illness, or been affected by a toxic buildup of some sub- stance or substances 2due to a lack of filtration in his tank for 27 days. "2's tank may have Another possibility is that the operating fi:lter in S.A.,r reduced the concentration of L-dopa below an effective level. E. Experimnt 4: Response to AC and pulsed magnetic fields in the T maze. 1. Conditioning of the sixty-cycle field. 2 The acrylic plastic T maze was prepared and the apparatus suitably modified to initially produce an alternatin- magnetic field and later a pulsed, but unidirectional field. A new line of investigation was then undertaken. We were now able to change the intensity of the mag- netic field over a considerable ran-e frcm-i 0 to over 50 gauss with a 2 simple adjustment of a variable AC ("Variac") transformer, or 0 - 20 gauss with the DC power source. The AC field was pulsed at the 60- cycle commercial frequency; the previously described circuit allowed the DC field to be pulsed from 0 - 1500 cps with no directional change. in the field. The chan,,es in frequency and intersity could be made 2 concurrently, although the complete range of intensities could not be achieved at every frequency. The first trials were run with a 60 cps AC mgretic field at 6 intensity levels (see Table 5). S.A.#3 was placed in the T maze, with the two magnets encircling one arm. The intensity of the magnetic field in that arm Fas var2ied by adjusting the voltage on the Variae. Freqt@ency was maintained at a constant 60 cps. At 41 gauss (50 volts), the fish was given 250 trials (on alternate trials, the field was turned on and off). 'When the field was on, each time the subject entered the experimental arm of the maze it was shocked briefly. With the field off, 2the S could enter and swim in the arm freely. It was hoped that in this way, i.e., by conditioning the S to avoid the field as an avers- ive stimulus by pairing it with shock, evidence could be obtained as to whether the fish was sensitive to the field. If candi tioning was achicved, then it would be definite that the S could perceive magnetic stim-ili, and its threshold for such stimuli determined by lowering the 22 intensity of the field. The first tests (250 trials) were made at 41 gauss. Aftc-.r 100 trials at this level, the fish never entered the arm with the field on, but swam into it when the field was off. Twenty trials were then attempted at 34, 36, 18@ 9 and 1 gauss, with similar positive results at the three higher setting2s. How- ever, at 9 and 1 gauss, behavior became inconsistent,. with @z!%e S re- sponding correctly about half the time. Table 5. Sixty--cps trials at intensities from 1 to 41 gauss. Intensity No. of trials Results 41 gatiss 250 Positive response 34 ga2uss 20 Positive response 26 gauss 20 Positive response 18 gauss 20 Positive response Partially positive 9 gauss 20 response ly PO,@ic2ive 1 gauss 20 sp-onse 2. Conditioning of variable-frequency fields. In each series of these trials the intensity of the magnetic field was held constant while frequency was varied by changes of 100 cps from 500 cps below the fish's (S.A.#3) own frequency to 500 cps above it (Table 6). A total of 1100 trials were run, 100 at eaci frequency. The method used was similar to-that of the pre- vioils experiment. That is, the field was turned on and off on al- ternate trials during each series of 100, and each time the fish went i2nto the arm of the maze with the field on it was shocked. When the field was off, no shock was used. The results in each case were negative; the fish did not learn to respond to the field as a noxious stimulus as we had expected on the basis of previous results, but tended to enter and stay in the field regardless of the shock. Thus, the field seemed to have son* posit2ive reinforce- ment value to the fish. This surprising outcome was checked in another series of 300 trials with S.A.#3, 100-at his own frequency (IC40 cps) and 100 each at 540 and 1540 cps. In these tests, however, the intensity of the field was increased to 15 gauss. The results were same; the S would not avoid the field whe9n paired with shock, but tend@@d to approach under all conditions. This interesting de- velopment led to the final and most important experinvnt in the pres- ent study, wherein the preference of the S's for the magnetic field was more fully explored. 23 Table 6. Conditioning with frequencies from 540 1540 cps at 5 gauss. Frequency (cps) No. of trials Results 1540 100 negative 1440 100 negative 1340 100 negative 2 1240 100 negntive 1140 100 negative 1040 (S''s own 100 negative frequency) 940 100 negative 840 2negative 740 100 negative 640 100 negative 540 100 negative 3. Approach response to pulsed fields with frequency and intensity varied. Up until this point, the evidence was somewhat contradictory and 2 .it was not sufficiently clear that the subjects were sensitive and re- sponsi-;e to magnetic fields. Ttierefore, on the basis of the positive evidence in the last experiment, it was decided to conduct a more comprehensive study of the fish's @reference for or approach tendency to the magnetic field. Tliie T maze was us2ed as before, with the magnets positioned on either side of one arm (Figure 5). Two series of trials were.run, one at 10 gauss and the other at 20 gauss. Each of the four speci- mens of Sternarchus albifrons was put in the maze, and the frequency was adjusted to the subject's own discharce rate, which, in these fish, was 700, 2885, 935, and 1,040 cps respectively. In subsequent trials the frequency was raised and lowered 100 and 200 cps above and below each subject's normal discharge rate at 26.80C. Thus, the experimental design involved changes along two continua, frequency and intensity (Table 7). Under each condition, the fish was placed into t2he maze, and the number of times it entered the experimental area in the arm between the magnetic coils during a period of 15 min- utes with the field off a-&id 15 minutes with the field on i;as recorded on a counter. Each fish was tested for its tendency to approach the field at five frequency levels ranging from 200 cps below to 200 cps 8 above its own frequency and at two intensity levels. A su=ary of the results are presented in Tables 8 and 9 and illustrated in Figure 9. 24 Table 7. Experirrental conditions for the approach-response experiment with a :-.ul,;ed magnetic field. 10 gauss 20 gauss S.A.1:1085 15 min on, 15 min off 15 min on, 15 min off S.A.2:900 +200 c2ps S.A.3:1240 S.A.4:1135 S.A.1:985 S.A.2:800 +100 cps S.A.3:1140 S.A.4:103 S.A.1:885 normal S.A.2:700 rate S.A.3:1040 S.A.4:935 S.A.1:7285 S.A.2:600 -100 cps S.A.3:940 S.A.4:835 S.A.1:685 S.A.2:500 -200 cps S.A.3"840 S.A.4:735 Table 8. Summa--y of results for the approach experiment, showing the number of times the subjects 2 entered the experimental area with the field on and off. S.A.1 S.A.2 S.A.3 S.A.4 10 20 10 20 10 20 10 20 gauss gauss gauss gauss gauss gauss gauss gauss total on 36 41 69 2 56 92 87 27 81 489 +200cps off 44 44 35 49 50 54 6 45 327 on 34 44 .49 36 59 74 53 78 427 +100cps off 21 17 33 29 42 42 53 52 289 ,6(srm. on 38 @8 34 43 27 14 19 23 236 2 -freg. off 36 28 26 21 29 17 1 11 16 184 on 38 46 44 61 52 50 60 59 410 -IOOCDS offl .25 34 28 32 46 27 39 39 270 oni 33 57 52 72 45 49 54 l@57@ 419 .:@c s@l -.,of2 f 144 44 41 1 41 54 36 40 1 39 1 339 As we can see in the total column, there was a very definite tendency for the Ss to enter the area between the magnets signi 'ficantly more times with the field on than,wlth the field off. For all fish and under all conditions, the experimental area was entered 1,981 times 2 with the field on, and 1,409 times with the field off. Means were cal- culated for the average number of times the fish entered the area with the field on and off under all frequencies and at the 10 and 20 gauss levels. The results are shown in Table 9. To test the significance of the difference between the combined means, a test was done with the following results: R=49.53 (field on mean) 7-35.23 (field off mean) B-14.30 (difference) EDi=-56-2- BDi2=14,956 N-40 Di-14.30 2_ E(y,7)2 Sd=10.35 4288 107.20 N 40 S 7.20 10.35 Sd 2 Off 10.35 1.66. VN- --I 6.25 t 14.30 = 8.61 We can thus reject the null hypothesis that the results occurred by chance at the .01 level, i.e.' we can be 99% confident that the ob- served difference was not due to chance. Table 9. Mean number of times the Ss entered the experimental area with the field on and off at 10 and 20 gauss and overall for al,l frequencies combined. 10 gauss 20 gauss combined Field on 45.75 53.30 49.53 Field off 35.15 35.30 35.23 In Figure 9, average difference scores were det2ermined by finding the difference between the number of times the Ss entered the experi- mental area with the field on and with it off, and dividing this number by 4 (the number of Ss). This was th'en plotted against the various fre- quencies tested. It can be seen that there are two definite peaks in the approach of the Ss to the field, at -100cps and in the re- 2 gion between +100 to +.200 cps. Surprisingly, there was a sharp drop at the subjects' own frequency, which is contrary to what we had expected, Previous reports had indicated that these fish are most sensitive at their own discharge frequency. After this analysis of the data, it was apparent that the fish were not responding to a pulsed magnetic 7field in the expected manner, i.e,, responses at the frequency of the fish at maze temperature were not the maximal responses observed, nor were the frequencies at which maximal responses were seen related in a periodic manner to the base (26.80C) frequency of the fish. 26 20-- 2.o 0 0 ........ 4 fa 41 4 J- 0 2 f efec'tri'c@l dischargel--,.-.-. Frequency of:ma'Snetic. f iolcis in-i6l'at3'.on to,fish-s frequency;o Figure 9. Changes in average difference scores with magnetic field frequency. Therefore, an additional series of trials were run extending the frequency 2range of the magnets. A field strength of 20 gauss was used throughout to maximize responses. The procedure'was-the same as before. The experimental fish was 'Placed in the maze, the lights were extinguished, and two minutes al- lowed to elapse before counting began. The number of times in a 15- minute period that the fish entered between the magnetic coils with 8 the magnets off was then recorded. With the electrical input to the magnet ard to the switch adjusted to produce 20 gauss and the desire'd frequency with the standardized pulse as shown -1@n Figure 5 for the mag- net longitudinal axis, the entries into the area between the magnets 27 GNETIC ONTAL tiAGNET FREQIIENCY IIORIZ LD. SCALE, 100 Hz. DIVISIONS. BASE SA I FISII FREQUENCY IN CENTER2 ed on performance a 20 Gauss field. 385 1385 SA 2 00 1500 co -SA 3 LU4U 1440 SA 4 3 85 935 0 in 15 minutes with the magnets on and off were counted. After each series of half hour trials, the fish was returned to his home tank, the n.,agnets readjusted if necessary to accommodate the frequency of the rext subject, and the process started aoain. In any given day, one 2fish would have a maximum of two half-hour trials, separated by at least one hour. The results are shown in Figure 10. Tnese per-formance curves were drawn by subtracting the number of entries durina each control run from the nurber of entries during the companion experimental run at each ma-net frequency.level. In actual numbers, the entries under cont-rol conditions ranged from 16 to 90 and under experimental condit tions ran-ed from 14 to 87 in a 15-minute period. Limitations in the equipment prevented the testing of performance at frequency levels of half and double that of the base frequency of ea@-,h fish. However, the range tested was adequate to show that sen- sitivity to a magnetic field in Sternarclius albifrons is vastly dif- ferent from that to an electric current. All of the data in previous reports with regard to response to magnets have been discussed in terms of the current generated in the fish by the magnetic fi2eld. This may be t-.ue, but these performance curves clearly indicate that the sensitivity to magnetism is more complex. The literature is in agreement that maximum sensitivity to applied current occurs at the fish's own frequency (Granath, 1967, Figure 11). The results reported here indicate that maximum sensi- 2 tivity to a magnetic field occurs at a point or points one to three hundred Hertz above and/or below the base frequency. What is most certainly indicated is that raxiiru,.n A.2 s@itivi@ty does not occur at the fish's own fre with the ma2netic field. so 2 7. 2 Z30 - 20 ;:Io 2 3 4 I 0 Fngou[Ncy (HZ) 2 Figure 11. Response spectrum to a uniform AC field. (After Cranath, 1967). If, as our data indicate, the fish respond to more than induced current in a pulsing magnetic field, there is still the problem of defining more clearly the stimuli to which they are responding. A crude model of a fish @as achieved by moving an inducti6on coil through the maze. Results'are reproduced in part in Figure 5. These wave 29 forms were induced with the axis of the test coil parallel to the axis of the riagnetic coils. With the a-is of the induction coil perpendic- ular to the axis of the magnetic coils,results were similar from the distal end of the test arm to the point 15 cm from the magnet center, but induced currents were weaker, 2 - 50 mV as op2posed to the 5 - 80 mV sho%4n on the diagram. However, from 15 cm, on toward the magnet center, the induced pulses became more roundcdand diminished in strength to about I gauss at the center of the magnetic coil. A behavior pattern observed in the fish indicated that the first model, though extremely crude, was better than2 the second. The fish, in moving from the distal end of the test arm would hesitate at about ttis 15 - 18 cm area, and then frequently continue to the magnet center in a rush, working their jaws and moving in an exciied manner. Interestingly, the induced cur- rent pulse in the-test coil assumes a wave form that is very like the fish's own di2scharge at this 15 - 18 cm point in our test apparatus, One other aspect of observed behavior toward the magnetic field is unexplained. When the fish chose the leg of the T at a point 0 to 10 cm from the intersection, they %iere observed to sometimes execute a forward roll, frequently two or three in succession with some2 degree of force. As can be seen from the diaaram, there appears to be no in- dividuality in this area of the field. This response was seen to some degree at all frequencies and-at 10 and 20 ga-ass, but seemed to be most common at those frequencies of maximum response to the magnet. A crude three-dimensional plot of the field shows2 it is cigar shaped. The fish were restricted to an area + 5 cm above and below the edge of the ci- gar-shaped field (23 cm in 'jiatreter). At this 10 cm point, the lines oi equal force would be essentially parallel to the long axis of the arm in the vertical plane, and cur-,.,ing toward the magnet center line in2 the horizontal plane. Perhaps it is this gradient to which the fish respond in this manner. F. Summary of results. (a) The rate of discharge in the electric field of Ster-narchus albi- frons is a positive function of temperature. In three subjects (S.A.1, 3, and 4) the change was + 50 cps for 1 degree C. The other specimen, 2 (S.A.2) varied + 15 cps per 1 degree C. (b) The fish showed no significant approach or avoidance behavior to- -ward a static (non-pulsed) magnetic field of 9 - 10 gauss in a Y maze. (e) A conditioning procedure in which elec'tric shock was paired with a static magnetic field of 9 - 10 gauss and 3k gauss in a Y maze in 0 order to establish an avoidance response was not successful. (d) In a study of the effects of drugs, Nerbut,il and levodopa (L. dopa) failed to alter the discharge patterns of the subjects' field, al- though L-dopa produced a more variable temperature-frequency relationship 30 'and some abnormal behavior. Pontocaine, however, modified the discharge frequency by depressing the rate sever6ly. (e) Conditioning trials were attempted with a 60-cycle AC magnetic field of I - 42 gauss paired with electric shock in a T maze. The fish learned @.o avoid the field at intensities of 34, 26, and 218 gauss, but the results were inconsistent at 9 and 1 gauss. (f) Additional conditioning trials were run in the T maze with a pulsed, unidirectional field at frequencies ranging from 540 to 1540 cps at 5 and 15 gauss in which the field was paired with shock, but the subject failed to learn to avoid the field. Instead, the fish showed 2 a tendency to stay in the field regardless of the shock it received. 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An electrical receptor in fishes. Nature, 1958, 131, 64-65. V. WHERE - 7ABO'ttATORY INDEX. The following is a list of laboratories,uuniversities0, and institutes where work is in progress. University of California Graduate School San Diego-LaJolla, California 92038 (Prinipal investigator: Th. Bullock) 34 Thp- City College of New York The City College Research Foundation Ne,w York, L%led York 10031 (Principal investigator: Frank J. Mandriota) Columbia University College of Physicians and Surgeons New-York, New York 10027 (Principal investigators: Arthur Karlin; David Nachmansohn) University of Connec2ticut Biological Sciences Group Storrs, Connecticut 06368 (Principal inves,tigator:.Tobias L. Schwartz) University of Connecticut Graduate School Storrs, Connecticut 06268 (Principal investigator: A.W. Wachtel) University of Ilaryland School of Yedicine Baltimore, Ilaryland 8 (Principal investigator:L. Holdman) Pennsylvania Hospital Philadelphia, Pennsylvania (Principal -'-nvestigator: Su=er I. Zacks) 35