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A method for simultaneously recording neural activity and rotation in the rat
Physiology & Behavior, Vol. 30, pp. 653-657. Pergamon Press Ltd., 1983. Printed in the U.S.A.
BRIEF COMMUNICATION
A Method for Simultaneously Recording Neural Activity and Rotation in the Rat J. S. M c K E N Z I E , P. W. E V E R E T T A N D L. J. D A L L Y D e p a r t m e n t o f Physiology, University o f Melbourne, Parkville, Victoria 3052, Australia R e c e i v e d 26 M a y 1982 McKENZIE, J. S., P. W. EVERETT AND L. J. DALLY. A method for simultaneously recording neural activity and rotation in the rat. PHYSIOL BEHAV 30(4) 653-657, 1983.--An apparatus is described which allows the simultaneous recording of neural activity and rotational behaviour in the rat. Rotational behaviour is measured, coded and stored on magnetic tape along with neural activity from chronically implanted electrodes, allowing off-line computer analysis. Rotational behaviour
Chronic recording
Neural activity
IN recent years many investigations of basal ganglia function have been made using the circling behaviour seen in rats following unilateral destruction of the dopaminergic nigrostriatal pathway and subsequent administration of dopaminergic stimulants (see [3] for review). The recording of neuronal activity in behaving animals, extensively employed in a number of other areas (e.g. [2]) has not previously been applied to investigations of the rotating preparation. It would, however, give direct information on neural mechanisms in the brain that are involved in producing this abnormal pattern of movement. The apparatus described here allows the simultaneous recording of neuronal activity and rotation. An important new feature of this method is the apparatus used for measuring and coding rotational movements. APPARATUS
General Description of the Apparatus Two pieces of Amphenol Micro-Rac plug strips glued together form the headpiece, a bridge between eight implanted fine-wire recording electrodes and a connector plug (Fig. la). The electrodes are constructed from 0.003 inch (0.076 mm) stainless steel wire with an insulating coat of teflon giving a total diameter of 0.0045 inches (0.11 mm) (Medwire No. 316). Electrodes are manufactured using a technique based on that described by Olds [2]. Gold plated female Amphenol Relia-Tac contacts are crimped onto the implanted electrodes once cemented in place, and inserted into the headpiece. The whole assembly is then firmly cemented to the skull (Fig. lb). A head amplifier is constructed from 8 F E T source followers ~mounted in a small aluminum box which plugs directly into the headpiece. The connector is assembled from Micro-Rac strips and male Relia-Tac pins. Signals from the head amplifier pass in a low resistance multiwire cable to an Air Precision 13 E E G multiple slip ring com-
mutator suspended above the animal and thence to a threechannel biological amplifier. The cable also transfers rotational movement of the animal to the commutator, where it is detected by the movement sensor (see below). Each channel of the amplifier has a gain selector switch offering the choice of 100× or 1000x amplification. High frequency (choice of 5, 7.5 or I0 KHz) and low frequency (choice of 100, 250 or 500 Hz) active filters are included in each channel. The filters have a roll-off of 12 db/octave. Rotational activity is measured by a movement sensor, a rotation analyser and a digital counter (Fig. 2), Signals from the rotation analyser are coded in a pulse mixer for storage on magnetic tape along with unitary or multiple unit neural activity recorded from implanted electrodes.
The Movement Sensor The movement sensor is attached to the top of the commutator (Fig. 3a) and generates four signals used by the rotation analyser to discriminate partial and full turns in either direction. A reflector disc, with 135° of arc removed, is attached to the axial shaft (originally designed for use in chronic infusions) protruding from the commutator. Fixed to the frame of the commutator and above the disc are four Optron Inc. OPB 706a infra-red reflective photocouplers mounted 90 ° apart. Each photo-coupler emits a beam of light which is reflected back by the disc to its photo-sensor. There is no reflectance if the gap lies beneath the photo-coupler. As the animal rotates in one direction, the light to the photocoupler is interrupted bi-sequentially (Figs. 3b and 3c). In the reverse direction the sequence of activation is reversed. There are eight distinct sensor activation modes for a 360 ° rotation, thus four sensors can distinguish eight 45 ° sectors.
Rotation Analyser and Counter Single sector and full turn signals in either direction are
Copyright © 1983 Pergamon Press Ltd.--O031-9384/83/040653-05503.00
654
McKENZIE ET AL.
j/-- to commut ator ---...~
I
I
AmpLifier
( Dim. = cm)
Z bend --/
(O)
Electrode (b)
FIG. 1. Headpiece and head amplifier, with connections to chronically-implanted electrodes.
produced by the rotation analyser from the sensor signals, and the number of each is counted and displayed by a four channel digital counter. The signals from the four sensors, after Schmitt trigger shaping, are re-encoded by an encoder/latch controller to briefly fix input data for transmission in a binary code to the sequence analyser. To de-
crease problems due to edge-jitter, the controller has an output which causes the input latch to hold data for 2 msec at every change of sector. It also drives a sensor position display, a circle of eight lamps numbered 0 to 7 on the panel of the instrument (Fig. 4b). The sequence analyser is an asynchronous finite-state machine [1] which, for each change of input, generates a short series of output changes dependent upon the previous states of the input. It monitors the sequence of rotation of the sector disc, and generates signals for either forward or reverse movements through sequential sectors. The signals appear as positive pulses on either the "forward" or "reverse" l/Sth turn output connectors for storage and further analysis. Two dual purpose up/down counters use these pulses to compute full 360° turns in both forward and reverse directions. In one mode a continuous sequence of eight sector transitions in one direction is required to produce a positive pulse on either the "forward" or "reverse" full turn output connectors, and any reversal during that sequence will reset the counter to zero. The start and end of the sequence will obviously be the sector which generates the eighth pulse. In the second mode, completion of a count of eight transitions in one direction, even if interrupted by reversals in direction, will generate a positive pulse at the relevant output connector. The counters cannot count below zero, so in the second mode the start and end of the completed circle will again be the sector which generates the eight pulse. In both modes the respective counter is reset to zero upon completion of a circle. A reset button clears the counters and temporarily fixes the start of a sequence. All four outputs from the rotation analyser are accumulated in four 5-digit counters each based around NonLinear-System Inc. PC4 event counters. A more complete description of the electronic circuits is contained in a paper in progress.
5
/
/ / / / / / / / /
/
/
FIG. 2. Experimental layout of equipment. (1) animal in bowl; (2) electrode cable; (3) movement sensor and commutator; (4) electrode signal cable; (5) rotation analyser with position display; (6) quad event counters; (7) pulse mixer; (8) 3-channel preamplifier; (9) 4-channel tape recorder.
RECORDING
NEURAL
ACTIVITY/ROTATION
IN RATS
655
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5
B
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A
~3~J
F I G . 3c
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l
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Rotation o f open sector from R e f 0
I
S e n s o r s activity D
0-45t 45090 ° 90°-135 ° 135°-180 ° 180°-225 ° 2250-270 ° 2700315 ° 315°-360/0 °
(a)
C
B
1 - 1 1 1 1 1 - -
Input code to Sequence A n a l y s e r
A
1 1 l I 1 -
-
Sector Number
0 1 2 3 4 5 6 7
1
c
b
a
0 0 0 1 1 1 1 0
0 1 1 1 1 0 0 0
1 1 0 0 1 I 0 0
FIG. 3. (a) M o v e m e n t s e n s o r and c o m m u t a t o r a s s e m b l y . (1) c o m m u t a t o r ; (2) m o v e m e n t s e n s o r housing; (3) notched disc on c o m m u t a t o r shaft; (4) printed-circuit with optoelectronic sensors; (5) s e n s o r output cable; (6) electrode cable to and from c o m m u t a t o r ; (7) support clamp. (b) Sensor relationship to slotted disc. (c) Sensor activity in relation to position o f slotted disc as s h o w n in b.
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Up/Down Counters & Control Logic
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FIG. 4. (a) Principle o f operation o f rotation encoding with the pulse mixer. (b) Block circuit diagram of rotation analyser.
M c K E N Z I E ET AL.
656
!
O
L
b
2Blab
4Blala
TIME
(SEC)
TIME
(SEC)
AMPHETAMINE
d
I
11"I
IPSIVERSIVE
ROTATION
o Q
21~e~
4~m~
FIG. 5. Records of single-unit impulses, unit firing-rate, and rotation rate of rats with unilateral 6-hydroxydopamine lesion of substantia nigra, (a) Bursting units of different amplitude fire at separate times in globus pallidus on lesioned side. (b) Steady firing of a unit in substantia nigra pars reticulata on side of lesion. (c) and (d) Unit firing-rate in globus paUidus of intact side, and rate of rotation towards lesioned side, before and after IP administration of dexamphetamine sulphate (3 mg/kg). (c) Unit rate on ordinate as mean impulses per second over 60 sec epochs (middle trace) with standard errors (top and bottom traces). (d) Rotation rate on ordinate as mean 45° turns per rain in constant direction over 60 sec epochs. Calibration: (a) 0.2 mV, 10 msec; (b) 0.5 mV, 10 msec.
Pulse Mixer The four outputs of the rotation analyser are encoded and summed into one signal suitable for recording on magnetic tape. Forward and reverse turns are differentiated by polarity, and 45° and 360° turns in the same direction are differentiated by amplitude. The signal output line carries a series of 0.5 msec positive or negative pulses in an amplitude ratio of 2:1 (refer to Fig. 4a).
Storage of Information The tape recorder used is a high quality domestic fourchannel reel-to-reel recorder. Output from the pulse mixer is stored in one channel and unit data from three electrodes at a time on the remaining channels. Each of the three amplifier channels can be switched to record activity against a common reference wire at any of the eight implanted electrodes. EXPERIMENTALPROCEDURE Adult male hooded rats are given a unilateral 6-hydroxydopamine lesion of the nigrostriatal pathway. Fourteen days are allowed for neuronal degeneration. Eight fine-wire recording electrodes are then implanted; 2 in the globus pallidus and 2 in the substantia nigra on each side of the brain are used at present. Ketalar anaesthesia is used for all surgery. The position of each of the eight electrodes is marked on the skull in ink, and eight small burr holes are drilled to allow insertion of the electrodes. Additional holes are drilled for anchoring screws and the earth electrode. The
neural activity on the electrodes is monitored as they are inserted under stereotaxic guidance. Each electrode is lowered to a position just above the target structure, then carefully advanced through it in search of suitable neural activity. When such activity has been isolated, the electrode is cemented in place with a drop of dental acrylic. When all eight electrodes have been suitably placed, they are connected to the headpiece as described above (Fig. 1). At least 4 days are allowed for recovery from implantation surgery, before experimental recording is begun. Extracellular recordings are made during a 15 minute pre-drug period and then during amphetamine (3 mg/kg IP) or apomorphine (l mg/kg SC) induced rotation, or with saline injected as control (0.9%, 0.25 ml IP), and stored along with coded rotational data on magnetic tape. The recordings are played back through an amplitude discriminator and the number of spikes falling within a given amplitude range is counted and stored on a computer floppy disc. The data on unit activity are displayed together with rotational activity by a digital plotter. Examples of unit discharge, firing rate and rotation rate are shown in Fig. 5. In one session only 3 electrodes can be recorded from at present due to limitations of our available equipment, making repeated trials necessary in order to obtain records from all 8 electrodes in parallel with behaviour; ideally, 8 channels of neural activity would be recorded simultaneously with rotation. At the completion of the series of trials a l0 p.A anodal current is passed down each electrode for l0 seconds to
R E C O R D I N G N E U R A L ACTIVITY/ROTATION IN RATS deposit iron at the tip. The animal is then perfused through the heart with saline followed by 10% potassium ferrocyanide in 10% formalin to form a prussian blue spot at each recording site. After further hardening in 10% formalin, the brain is removed from the skull, cut in frozen sections and stained on the slide with cresyl violet. The recording sites are then verified. RESULTS AND DISCUSSION The apparatus has been used successfully in this laboratory for over 12 months. The pseudo-peak-to-peak instrumentation noise of the system (measured with shorted input) is 12/zV for the amplifier and FET, and 15/~V for the amplifier, F E T and electrode submerged in 0.9% saline. With the electrode implanted in the animal, biological noise increases the unresolved background to 50-60/zV. A signalto-noise ratio of 2 or 3 to 1 is achieved for most units, but it is
657 possible to record single units up to several mV in amplitude (Fig. 5). The apparatus does not remove the necessity for visual observation of the animal in order to discriminate different motor patterns of circling. F o r example, brief changes in direction, such as may occur in rearing or headswinging, may produce 45 ° sector impulses. This affects only the 45 ° counts, not the 360 ° count, and then only when the orientation hovers around a division between two 45 ° sectors. The electrode connector with cement (2.5 g), the F E T input head and recording cable do not present a significant burden to a rat's movement since their combined weight is almost entirely supported by the slip-ring assembly. Because of the low friction and inertia of the commutator, there is no significant cable torsion. The unimpended rotation of the rats in the circular enclosure is not observably different from that of other treated rats free of implants, connectors and cable.
REFERENCES
1. Fletcher, W. I. Asynchronous finite-state machines. In: An Engineering Approach to Digital Design. New Jersey: Prentice-Hall, 1980, pp. 649-717.
2. Olds, J. Multiple unit recordings from behaving rats. In: Bioelectric Recording Techniques, Part A: Cellular Processes and Brain Potentials, edited by R. S. Thompson and M. M. Patter-
son. New York: Academic Press, 1973, pp. 165-198. 3. Pycock, C. J. Turning behavior in animals. Neuroscience 5: 461-514, 1980.
BRIEF COMMUNICATION
A Method for Simultaneously Recording Neural Activity and Rotation in the Rat J. S. M c K E N Z I E , P. W. E V E R E T T A N D L. J. D A L L Y D e p a r t m e n t o f Physiology, University o f Melbourne, Parkville, Victoria 3052, Australia R e c e i v e d 26 M a y 1982 McKENZIE, J. S., P. W. EVERETT AND L. J. DALLY. A method for simultaneously recording neural activity and rotation in the rat. PHYSIOL BEHAV 30(4) 653-657, 1983.--An apparatus is described which allows the simultaneous recording of neural activity and rotational behaviour in the rat. Rotational behaviour is measured, coded and stored on magnetic tape along with neural activity from chronically implanted electrodes, allowing off-line computer analysis. Rotational behaviour
Chronic recording
Neural activity
IN recent years many investigations of basal ganglia function have been made using the circling behaviour seen in rats following unilateral destruction of the dopaminergic nigrostriatal pathway and subsequent administration of dopaminergic stimulants (see [3] for review). The recording of neuronal activity in behaving animals, extensively employed in a number of other areas (e.g. [2]) has not previously been applied to investigations of the rotating preparation. It would, however, give direct information on neural mechanisms in the brain that are involved in producing this abnormal pattern of movement. The apparatus described here allows the simultaneous recording of neuronal activity and rotation. An important new feature of this method is the apparatus used for measuring and coding rotational movements. APPARATUS
General Description of the Apparatus Two pieces of Amphenol Micro-Rac plug strips glued together form the headpiece, a bridge between eight implanted fine-wire recording electrodes and a connector plug (Fig. la). The electrodes are constructed from 0.003 inch (0.076 mm) stainless steel wire with an insulating coat of teflon giving a total diameter of 0.0045 inches (0.11 mm) (Medwire No. 316). Electrodes are manufactured using a technique based on that described by Olds [2]. Gold plated female Amphenol Relia-Tac contacts are crimped onto the implanted electrodes once cemented in place, and inserted into the headpiece. The whole assembly is then firmly cemented to the skull (Fig. lb). A head amplifier is constructed from 8 F E T source followers ~mounted in a small aluminum box which plugs directly into the headpiece. The connector is assembled from Micro-Rac strips and male Relia-Tac pins. Signals from the head amplifier pass in a low resistance multiwire cable to an Air Precision 13 E E G multiple slip ring com-
mutator suspended above the animal and thence to a threechannel biological amplifier. The cable also transfers rotational movement of the animal to the commutator, where it is detected by the movement sensor (see below). Each channel of the amplifier has a gain selector switch offering the choice of 100× or 1000x amplification. High frequency (choice of 5, 7.5 or I0 KHz) and low frequency (choice of 100, 250 or 500 Hz) active filters are included in each channel. The filters have a roll-off of 12 db/octave. Rotational activity is measured by a movement sensor, a rotation analyser and a digital counter (Fig. 2), Signals from the rotation analyser are coded in a pulse mixer for storage on magnetic tape along with unitary or multiple unit neural activity recorded from implanted electrodes.
The Movement Sensor The movement sensor is attached to the top of the commutator (Fig. 3a) and generates four signals used by the rotation analyser to discriminate partial and full turns in either direction. A reflector disc, with 135° of arc removed, is attached to the axial shaft (originally designed for use in chronic infusions) protruding from the commutator. Fixed to the frame of the commutator and above the disc are four Optron Inc. OPB 706a infra-red reflective photocouplers mounted 90 ° apart. Each photo-coupler emits a beam of light which is reflected back by the disc to its photo-sensor. There is no reflectance if the gap lies beneath the photo-coupler. As the animal rotates in one direction, the light to the photocoupler is interrupted bi-sequentially (Figs. 3b and 3c). In the reverse direction the sequence of activation is reversed. There are eight distinct sensor activation modes for a 360 ° rotation, thus four sensors can distinguish eight 45 ° sectors.
Rotation Analyser and Counter Single sector and full turn signals in either direction are
Copyright © 1983 Pergamon Press Ltd.--O031-9384/83/040653-05503.00
654
McKENZIE ET AL.
j/-- to commut ator ---...~
I
I
AmpLifier
( Dim. = cm)
Z bend --/
(O)
Electrode (b)
FIG. 1. Headpiece and head amplifier, with connections to chronically-implanted electrodes.
produced by the rotation analyser from the sensor signals, and the number of each is counted and displayed by a four channel digital counter. The signals from the four sensors, after Schmitt trigger shaping, are re-encoded by an encoder/latch controller to briefly fix input data for transmission in a binary code to the sequence analyser. To de-
crease problems due to edge-jitter, the controller has an output which causes the input latch to hold data for 2 msec at every change of sector. It also drives a sensor position display, a circle of eight lamps numbered 0 to 7 on the panel of the instrument (Fig. 4b). The sequence analyser is an asynchronous finite-state machine [1] which, for each change of input, generates a short series of output changes dependent upon the previous states of the input. It monitors the sequence of rotation of the sector disc, and generates signals for either forward or reverse movements through sequential sectors. The signals appear as positive pulses on either the "forward" or "reverse" l/Sth turn output connectors for storage and further analysis. Two dual purpose up/down counters use these pulses to compute full 360° turns in both forward and reverse directions. In one mode a continuous sequence of eight sector transitions in one direction is required to produce a positive pulse on either the "forward" or "reverse" full turn output connectors, and any reversal during that sequence will reset the counter to zero. The start and end of the sequence will obviously be the sector which generates the eighth pulse. In the second mode, completion of a count of eight transitions in one direction, even if interrupted by reversals in direction, will generate a positive pulse at the relevant output connector. The counters cannot count below zero, so in the second mode the start and end of the completed circle will again be the sector which generates the eight pulse. In both modes the respective counter is reset to zero upon completion of a circle. A reset button clears the counters and temporarily fixes the start of a sequence. All four outputs from the rotation analyser are accumulated in four 5-digit counters each based around NonLinear-System Inc. PC4 event counters. A more complete description of the electronic circuits is contained in a paper in progress.
5
/
/ / / / / / / / /
/
/
FIG. 2. Experimental layout of equipment. (1) animal in bowl; (2) electrode cable; (3) movement sensor and commutator; (4) electrode signal cable; (5) rotation analyser with position display; (6) quad event counters; (7) pulse mixer; (8) 3-channel preamplifier; (9) 4-channel tape recorder.
RECORDING
NEURAL
ACTIVITY/ROTATION
IN RATS
655
C
5
B
D
e ~
o)
A
~3~J
F I G . 3c
'
l
'
\
Rotation o f open sector from R e f 0
I
S e n s o r s activity D
0-45t 45090 ° 90°-135 ° 135°-180 ° 180°-225 ° 2250-270 ° 2700315 ° 315°-360/0 °
(a)
C
B
1 - 1 1 1 1 1 - -
Input code to Sequence A n a l y s e r
A
1 1 l I 1 -
-
Sector Number
0 1 2 3 4 5 6 7
1
c
b
a
0 0 0 1 1 1 1 0
0 1 1 1 1 0 0 0
1 1 0 0 1 I 0 0
FIG. 3. (a) M o v e m e n t s e n s o r and c o m m u t a t o r a s s e m b l y . (1) c o m m u t a t o r ; (2) m o v e m e n t s e n s o r housing; (3) notched disc on c o m m u t a t o r shaft; (4) printed-circuit with optoelectronic sensors; (5) s e n s o r output cable; (6) electrode cable to and from c o m m u t a t o r ; (7) support clamp. (b) Sensor relationship to slotted disc. (c) Sensor activity in relation to position o f slotted disc as s h o w n in b.
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(a)
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.
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.
.
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I•
Reset
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_ J
Up/Down Counters & Control Logic
/
(b)
FIG. 4. (a) Principle o f operation o f rotation encoding with the pulse mixer. (b) Block circuit diagram of rotation analyser.
M c K E N Z I E ET AL.
656
!
O
L
b
2Blab
4Blala
TIME
(SEC)
TIME
(SEC)
AMPHETAMINE
d
I
11"I
IPSIVERSIVE
ROTATION
o Q
21~e~
4~m~
FIG. 5. Records of single-unit impulses, unit firing-rate, and rotation rate of rats with unilateral 6-hydroxydopamine lesion of substantia nigra, (a) Bursting units of different amplitude fire at separate times in globus pallidus on lesioned side. (b) Steady firing of a unit in substantia nigra pars reticulata on side of lesion. (c) and (d) Unit firing-rate in globus paUidus of intact side, and rate of rotation towards lesioned side, before and after IP administration of dexamphetamine sulphate (3 mg/kg). (c) Unit rate on ordinate as mean impulses per second over 60 sec epochs (middle trace) with standard errors (top and bottom traces). (d) Rotation rate on ordinate as mean 45° turns per rain in constant direction over 60 sec epochs. Calibration: (a) 0.2 mV, 10 msec; (b) 0.5 mV, 10 msec.
Pulse Mixer The four outputs of the rotation analyser are encoded and summed into one signal suitable for recording on magnetic tape. Forward and reverse turns are differentiated by polarity, and 45° and 360° turns in the same direction are differentiated by amplitude. The signal output line carries a series of 0.5 msec positive or negative pulses in an amplitude ratio of 2:1 (refer to Fig. 4a).
Storage of Information The tape recorder used is a high quality domestic fourchannel reel-to-reel recorder. Output from the pulse mixer is stored in one channel and unit data from three electrodes at a time on the remaining channels. Each of the three amplifier channels can be switched to record activity against a common reference wire at any of the eight implanted electrodes. EXPERIMENTALPROCEDURE Adult male hooded rats are given a unilateral 6-hydroxydopamine lesion of the nigrostriatal pathway. Fourteen days are allowed for neuronal degeneration. Eight fine-wire recording electrodes are then implanted; 2 in the globus pallidus and 2 in the substantia nigra on each side of the brain are used at present. Ketalar anaesthesia is used for all surgery. The position of each of the eight electrodes is marked on the skull in ink, and eight small burr holes are drilled to allow insertion of the electrodes. Additional holes are drilled for anchoring screws and the earth electrode. The
neural activity on the electrodes is monitored as they are inserted under stereotaxic guidance. Each electrode is lowered to a position just above the target structure, then carefully advanced through it in search of suitable neural activity. When such activity has been isolated, the electrode is cemented in place with a drop of dental acrylic. When all eight electrodes have been suitably placed, they are connected to the headpiece as described above (Fig. 1). At least 4 days are allowed for recovery from implantation surgery, before experimental recording is begun. Extracellular recordings are made during a 15 minute pre-drug period and then during amphetamine (3 mg/kg IP) or apomorphine (l mg/kg SC) induced rotation, or with saline injected as control (0.9%, 0.25 ml IP), and stored along with coded rotational data on magnetic tape. The recordings are played back through an amplitude discriminator and the number of spikes falling within a given amplitude range is counted and stored on a computer floppy disc. The data on unit activity are displayed together with rotational activity by a digital plotter. Examples of unit discharge, firing rate and rotation rate are shown in Fig. 5. In one session only 3 electrodes can be recorded from at present due to limitations of our available equipment, making repeated trials necessary in order to obtain records from all 8 electrodes in parallel with behaviour; ideally, 8 channels of neural activity would be recorded simultaneously with rotation. At the completion of the series of trials a l0 p.A anodal current is passed down each electrode for l0 seconds to
R E C O R D I N G N E U R A L ACTIVITY/ROTATION IN RATS deposit iron at the tip. The animal is then perfused through the heart with saline followed by 10% potassium ferrocyanide in 10% formalin to form a prussian blue spot at each recording site. After further hardening in 10% formalin, the brain is removed from the skull, cut in frozen sections and stained on the slide with cresyl violet. The recording sites are then verified. RESULTS AND DISCUSSION The apparatus has been used successfully in this laboratory for over 12 months. The pseudo-peak-to-peak instrumentation noise of the system (measured with shorted input) is 12/zV for the amplifier and FET, and 15/~V for the amplifier, F E T and electrode submerged in 0.9% saline. With the electrode implanted in the animal, biological noise increases the unresolved background to 50-60/zV. A signalto-noise ratio of 2 or 3 to 1 is achieved for most units, but it is
657 possible to record single units up to several mV in amplitude (Fig. 5). The apparatus does not remove the necessity for visual observation of the animal in order to discriminate different motor patterns of circling. F o r example, brief changes in direction, such as may occur in rearing or headswinging, may produce 45 ° sector impulses. This affects only the 45 ° counts, not the 360 ° count, and then only when the orientation hovers around a division between two 45 ° sectors. The electrode connector with cement (2.5 g), the F E T input head and recording cable do not present a significant burden to a rat's movement since their combined weight is almost entirely supported by the slip-ring assembly. Because of the low friction and inertia of the commutator, there is no significant cable torsion. The unimpended rotation of the rats in the circular enclosure is not observably different from that of other treated rats free of implants, connectors and cable.
REFERENCES
1. Fletcher, W. I. Asynchronous finite-state machines. In: An Engineering Approach to Digital Design. New Jersey: Prentice-Hall, 1980, pp. 649-717.
2. Olds, J. Multiple unit recordings from behaving rats. In: Bioelectric Recording Techniques, Part A: Cellular Processes and Brain Potentials, edited by R. S. Thompson and M. M. Patter-
son. New York: Academic Press, 1973, pp. 165-198. 3. Pycock, C. J. Turning behavior in animals. Neuroscience 5: 461-514, 1980.