J. Physiol. (1977), 264, pp. 1-16 With 1 plate and 5 text-fgure Printed in Great Britain
1
INHIBITION OF SPINOCERVICAL TRACT DISCHARGES FROM LOCALIZED AREAS OF THE SENSORIMOTOR CORTEX IN THE CAT
By A. G. BROWN, J. D. COULTER,* P. K. ROSEt A. D. SHORT AND P. J. SNOWt From the Department of Physiology, Royal (Dick) School of Veterinary Studies, University of Edinburgh, Edinburgh EH9 1QH
(Received 23 February 1976) SUMMARY
1. Intracortical microstimulation (IJMS) was applied within the sensorimotor cortex of cats anaesthetized with chloralose. 2. The effects of the ICMS were examined on the number of impulses in spinocervical tract (SCT) cells (recorded extracellularly in the contralateral lumbosacral spinal cord) evoked by peripheral stimulation. 3. Inhibition of SCT discharges was produced by ICMS in two distinct regions of the sensorimotor cortex. 4. One inhibitory region was in part of cytoarchitectonic area 4y in the upper bank of the cruciate sulcus. It sometimes extended caudally into area 4&, medially into area 3a and/or rostrally into the part of area 4y on the caudal lip of the cruciate sulcus. 5. The other inhibitory region was in the medial part of the posterior sigmoid gyrus and included parts of areas 3a, 3b, 1, 5a and 5b. 6. Most inhibitory sites were in cortical layers III, V and VI. 7. No regions were found in which ICMS consistently caused facilitation of SCT discharges. INTRODUCTION
Electrical stimulation of the surface of the sensorimotor cortex in the cat leads to primary afferent depolarization of cutaneous nerve terminals in the spinal cord (Carpenter, Lundberg & Norrsell, 1963; Andersen, Eccles & Sears, 1964c) through both pyramidal and extrapyramidal pathways (Hongo & Jankowska, 1967). The sensorimotor cortex also affects transmission through the dorsal column nuclei (Towe & Jabbur, 1961), in part * Fellow of the Foundations Fund for Research in Psychiatry. Present address: Marine Biomedical Institute and Departments of Physiology and Biophysics and Psychiatry, University of Texas Medical Branch, Galveston, U.S.A. t Post-doctoral Fellows of the Canadian M.R.C.
I-2
2 2 at least
~~~A. G. BROWN AND OTHERS
through presynaptic inhibitory action (Andersen, Eccies, Schmidt & Yokata, 1964b; Andersen, Eccles, Oshima & Schmidt, 1964a) although excitatory effects are observed as well as inhibitory ones (Jabbur & Towe, 1961; Gordon & Jukes, 1964; Levitt, Carreras, Liu & Chambers, 1964). Similar effects have been observed in spinocerebellar pathways (for review see Oscarsson, 1973) and in monkey dorsal column nuclei (Harris, Jabbur, Morse & Towe, 1965) and spinothalamic tract (Coulter, Maunz & Willis, 1974). The spinocervical tract (SOT) of the cat is also under inhibitory control from the sensorimotor cortex. Wall (1967) and Fetz (1968) showed, in decerebrate cats, that dorsal horn neurones, some of which give rise to the SOT, may be inhibited or excited by stimulation of the medullary pyramids, although Lundberg, Norrsell & Voorhoeve (1963) had observed little effect on the SOT upon stimulation of the surface of the sensorimotor cortex in barbiturate-anaesthetized animals. More recently Brown & Short (1974) have shown in chloralose-anaesthetized cats that transmission through the hind limb SOT may be inhibited easily and consistently by electrical stimulation of the sensorimotor cortex, most effectively from the hind-limb area of the first somatic sensory cortex, with a time course similar to that of primary afferent depolarization. The present experiments were designed to locate more precisely the cortical cytoarchitectonic areas from which inhibition could be produced. We used the intracortical microstimulation technique developed by Asanuma and his colleagues (Stoney, Thompson & Asanuma, 1968). A preliminary account of this work has been published (Brown, Coulter, Rose, Short & Snow, 1976). METHODS Fifteen cats (2 1-3-0 kg weight) were used. During the initial surgery the anaesthetic was 2-4 % halothane in 50 % NO2, 50% 02. At least 2 h before recording was begun this anaesthetic was discontinued and the experiment continued under chloralose anaesthesia (70 mg.- kg-" ixv.). The animals were paralysed with gallamine triethiodide, 40 mg i v. initially and 20 mng h'1 subsequently, and artificially respired. After tracheal, venous and arterial cannulations the sural and medial plantar nerves were exposed in both hind limbs. The spinal cord was exposed at upper cervical and lumbosacral levels and covered with liquid paraffin B.Px. at 370 C. The sensorimotor cortex contralateral to the lumbo-sacral recording site was exposed and covered with 50 % liquid paraffin B.P. 50 % petroleum jelly B.P. at 370 C. An enlarged photograph of the cortex was available during the recording session and was used to record the entry points of the stimulating micro-electrode into the cortex. A bilateral pneumothorax was performed. Rectal temperature was maintained close to 380 C with a thermostatically controlled electrical blanket on the ventral body surface. Arterial blood pressure was monitored and the end-tidal CO2 maintained at 3-5-4-0 % by adjusting the stroke volume of the respiratory pump. The
CORTICAL INHIBITION OF THE SOT
3
pump volume was adjusted so that the pupils remained constricted and this occurred within the above end-tidal C02 ranges. Extracellular recordings were made from SCT cells using glass micro-pipette electrodes filled either with 4 M-NaCl (resistance 2-5 Mil) or with 5 % Procion Yellow M4RS in distilled water (resistance about 10 Mil). Methods for identifying the SCT have been described previously and included antidromic activation from the dorsolateral funiculus at C 3, with collision of ortho- and antidromic impulses, and absence of antidromic activation from C 1. Responses were evoked in SCT cells by electrical stimulation of ipsilateral cutaneous nerves (0-2 ms single shocks) or by electrical stimulation of cutaneous receptive fields through a pair of tungsten wire electrodes inserted into the skin. The peripheral stimuli were adjusted until the SCT cell under study responded with 4-20 impulses. Units were discarded if the variability in the number of impulses was too great. Once an SCT cell had been isolated and its receptive field characterized, intracortical cathodal microstimulation was performed using tungsten in glass microelectrodes (Stoney et al. 1968) with exposed tungsten tips of 15-30 /stm. The anode was a silver wire buried in the temporal muscle. Stimulating current pulses were measured as the voltage drop across a 1 kQl series resistor between the isolated stimulator and the silver anode. After preliminary trials the cortical stimulus was fixed at 3 shocks, 0-2 ms duration at 400 Hz. The conditioning-testing interval was fixed at 40 ms which is at about the time of maximal inhibitory action elicited by stimulation of the cortical surface (Brown & Short, 1974). Two series of experiments were performed. In the first series the stimulating micro-electrode was driven through the cortex in steps of 200 Fzm until inhibitory effects were seen and then in 100 /um steps through inhibitory regions using cortical stimuli of 50 or 25 VIA strength. Once inhibition was observed the cortical shocks were reduced to 10 and 5 /%A in attempts to determine the lowest strengths which would affect transmission through the SCT. Several (up to six) tracks were made in this series, usually all close to each other (500 Fm) in the transverse and sagittal directions. In the second series as much of the sensorimotor cortex as possible behind the position of the cruciate sulcus at the surface was searched with a set of electrode tracks 1-0-1-5 mm apart in the sagittal and coronal directions in attempts to surround all inhibitory areas with negative tracks. For these experiments the stimulating current was fixed at 25 /zA and the stimulation sites were every 250 Fm in depth. In both series of experiments, for each stimulation site ten unconditioned and ten conditioned responses were recorded. Initially the ten control responses were recorded before the ten conditioned responses. In the last four experiments, all in the second series, the control and conditioned responses were alternated. The recorded impulses from the SCT cell were converted to standard pulses which were counted on a counter-timer. The number of impulses in unconditioned and conditioned responses were compared using Student's t test (grouped or paired t tests where appropriate; two-tailed tests, P < 0 05 accepted as significant; the variance of the control and conditioned responses were assumed equal). Responses were also recorded on tape for subsequent analysis. After the completion of the last stimulation track, in the second series of experiments, electrolytic lesions were made by passing 10 /FA for 10 a (electrode negative) at known distances apart in the last track. In all experiments the stimulating electrode was left at its deepest position in the last track. The brain was perfused with normal saline followed by 10 % formol-saline. The next day the block of brain containing the electrode tracks was removed, the rostral and caudal surfaces of the block being cut in the coronal plane of the electrode tracks. After further
4
A. G. BROWN AND OTHERS
fixation in formol-saline four of the first series of brains were cut at 25 jsm (frozen sections) and four of the first series and all the second series were cut at 15 jam after paraffin wax embedding. All sections were cut in the coronal plane and stained with Cresyl Violet. The stimulating tracks were reconstructed. Cytoarchitectonic areas of the sensorimotor cortex were identified in each individual brain by more than one of us, and it should be noted that the criteria of Hassler & Muhs-Clement (1964) were used, not their published illustrations. We found their descriptions to be accurate. Nomenclature. In this paper the term area means a cytoarchitectonic area. Site means the position of the intracortical electrode tip during microstimulation. An inhibitory 8ite is a site from which statistically significant inhibition of an SCT cell was elicited. A negative 8ite is a site from which no significant inhibition was elicited. An inhibitory region is a group of two or more inhibitory sites adjacent in the vertical or horizontal direction without intervening negative sites. B
A
C
50
~40 r-30 0 20 C
10
0
5 10
25 4uA
50
10 i
25
uA
50
5 10
25
4uA
Text-fig. 1. The relationship between the amount of inhibition of SCT units and strength of intracortical microstimulation current. A shows results when three different current strengths produced significant inhibition, B and C show results when 2 out of 3 current strengths produced significant inhibition. The relationships are non-linear with a reduction in slope as current is increased. Each curve represents data obtained from a single SCT unit by stimulation of a single cortical site. The sixteen curves are from sixteen cortical sites and seven SCT units. Circles: significant inhibition (P < 0.05). Triangles: non-significant values. RESULTS
The aim of the first series of eight experiments was to find those regions of cortex from which inhibitory actions could be elicited at minimal current strengths and initially currents of 50 #uA were used to search for these regions. The electrode was driven through the cortex and conditioning shocks given every 200 Mum until inhibition was observed. Inhibitory effects were usually observed suddenly at a particular electrode position and went away equally abruptly as the electrode passed through the inhibitory region. When clear inhibition was obtained with 50 ,sA currents then the electrode was moved in steps of 100 Aim and currents of 25, 10
5 CORTICAL INHIBITION OF THE SOT and sometimes 5 1sA were used to determine the minimal current necessary for eliciting inhibition. The amount of inhibition (%) produced by histologically confirmed intracortical micro-stimulation, 50 pA or less, was considerably smaller than that produced by surface stimulation in earlier experiments (Brown & Short, 1974). Both 50 and 25 aA produced significant inhibition at twenty-one cortical sites. At these sites 50 pA gave an average of 32-3 % inhibition ( ± 9 7, S.D.) while 25 pA gave an average of 26 3 % inhibition (± 10-7, S.D.). At twelve cortical sites where both 25 and 10 /tA produced significant inhibition the average amounts of inhibition were 27-5 (± 9 5 %, S.D.) and 20.7 ( ± 7 0 %, S.D.), respectively. Text-fig. 1 shows percentage inhibition plotted as a function of stimulus current for all sixteen sites, obtained from 7 SCT units, where significant inhibition was elicited by three currents (Text-fig. 1 A) or by two of three currents (Text-fig. 1 B, C). It is evident that the relationship is non-linear, with a reduction in slope as current is increased. The number of sites giving rise to significant inhibition was greater with larger currents. In general 25 ,uA produced smaller inhibitory regions than 50 ,LA and in turn 10 and 5 ,uA produced even smaller ones. The positions giving maximal inhibition with the smaller currents were always similar to those giving maximal inhibition with the larger currents. Of forty-one cortical sites at which significant inhibition was produced with either 50 or 25 flA, the 50 ,jA stimulus was effective at thirty-seven (90 %) of them and the 25 pA at twenty-four (59 %). The 25 1tA stimulus was effective at four sites where 50 /tA did not produce significant inhibition. More generally, the 50 pA stimulus revealed six inhibitory regions, in four brains, which were.also revealed by the 25 ,tA stimulus. Similarly, of sixty-four sites at which significant inhibition was produced with either 25 or 10 #uA, the 25 pA stimulus was effective at fifty-nine (92 %) and the 10 pA at seventeen (25 %). There were five sites at which only the 10 #uA stimulus produced significant effects. Although eleven distinct regions were revealed using 25 #uA only six of these were revealed using 10 #uA. A further reduction of current strength to 5 #uA led to significant inhibition at only one of these six regions. Location of inhibitory regions in cortical cytoarchitectonic areas. In the first series of experiments none of the cortical inhibitory regions was entirely surrounded by negative sites. The reconstructed brains did not allow an estimate to be made of the extent and position of each inhibitory region. In mapping the cortex in three dimensions a compromise must be made between the conflicting aims of precise localization of inhibitory regions and coverage of a large volume of cortex. Experience gained in the first series suggested that a grid of tracks about 1 mm apart was the finest
A. G. BROWN AND OTHERS which would permit an extensive search within the 12 h or so for which a single SCT cell could be recorded. In the second series of seven experiments (a single SCT cell was recorded in each) the strength of shock was fixed at 25 ,#A at sites 250 ,tm apart within tracks, since this had not missed any inhibitory regions detected by larger currents or smaller steps. Using these parameters cortical sites which produced significant inhibition nearly always had at least one other similar site in one neighbouring track (see Text-figs. 2-4 and P1. 1). Only 6
2.1mm
A
c
Inhibitio
P
1
INHIBITION OF SPINOCERVICAL TRACT DISCHARGES FROM LOCALIZED AREAS OF THE SENSORIMOTOR CORTEX IN THE CAT
By A. G. BROWN, J. D. COULTER,* P. K. ROSEt A. D. SHORT AND P. J. SNOWt From the Department of Physiology, Royal (Dick) School of Veterinary Studies, University of Edinburgh, Edinburgh EH9 1QH
(Received 23 February 1976) SUMMARY
1. Intracortical microstimulation (IJMS) was applied within the sensorimotor cortex of cats anaesthetized with chloralose. 2. The effects of the ICMS were examined on the number of impulses in spinocervical tract (SCT) cells (recorded extracellularly in the contralateral lumbosacral spinal cord) evoked by peripheral stimulation. 3. Inhibition of SCT discharges was produced by ICMS in two distinct regions of the sensorimotor cortex. 4. One inhibitory region was in part of cytoarchitectonic area 4y in the upper bank of the cruciate sulcus. It sometimes extended caudally into area 4&, medially into area 3a and/or rostrally into the part of area 4y on the caudal lip of the cruciate sulcus. 5. The other inhibitory region was in the medial part of the posterior sigmoid gyrus and included parts of areas 3a, 3b, 1, 5a and 5b. 6. Most inhibitory sites were in cortical layers III, V and VI. 7. No regions were found in which ICMS consistently caused facilitation of SCT discharges. INTRODUCTION
Electrical stimulation of the surface of the sensorimotor cortex in the cat leads to primary afferent depolarization of cutaneous nerve terminals in the spinal cord (Carpenter, Lundberg & Norrsell, 1963; Andersen, Eccles & Sears, 1964c) through both pyramidal and extrapyramidal pathways (Hongo & Jankowska, 1967). The sensorimotor cortex also affects transmission through the dorsal column nuclei (Towe & Jabbur, 1961), in part * Fellow of the Foundations Fund for Research in Psychiatry. Present address: Marine Biomedical Institute and Departments of Physiology and Biophysics and Psychiatry, University of Texas Medical Branch, Galveston, U.S.A. t Post-doctoral Fellows of the Canadian M.R.C.
I-2
2 2 at least
~~~A. G. BROWN AND OTHERS
through presynaptic inhibitory action (Andersen, Eccies, Schmidt & Yokata, 1964b; Andersen, Eccles, Oshima & Schmidt, 1964a) although excitatory effects are observed as well as inhibitory ones (Jabbur & Towe, 1961; Gordon & Jukes, 1964; Levitt, Carreras, Liu & Chambers, 1964). Similar effects have been observed in spinocerebellar pathways (for review see Oscarsson, 1973) and in monkey dorsal column nuclei (Harris, Jabbur, Morse & Towe, 1965) and spinothalamic tract (Coulter, Maunz & Willis, 1974). The spinocervical tract (SOT) of the cat is also under inhibitory control from the sensorimotor cortex. Wall (1967) and Fetz (1968) showed, in decerebrate cats, that dorsal horn neurones, some of which give rise to the SOT, may be inhibited or excited by stimulation of the medullary pyramids, although Lundberg, Norrsell & Voorhoeve (1963) had observed little effect on the SOT upon stimulation of the surface of the sensorimotor cortex in barbiturate-anaesthetized animals. More recently Brown & Short (1974) have shown in chloralose-anaesthetized cats that transmission through the hind limb SOT may be inhibited easily and consistently by electrical stimulation of the sensorimotor cortex, most effectively from the hind-limb area of the first somatic sensory cortex, with a time course similar to that of primary afferent depolarization. The present experiments were designed to locate more precisely the cortical cytoarchitectonic areas from which inhibition could be produced. We used the intracortical microstimulation technique developed by Asanuma and his colleagues (Stoney, Thompson & Asanuma, 1968). A preliminary account of this work has been published (Brown, Coulter, Rose, Short & Snow, 1976). METHODS Fifteen cats (2 1-3-0 kg weight) were used. During the initial surgery the anaesthetic was 2-4 % halothane in 50 % NO2, 50% 02. At least 2 h before recording was begun this anaesthetic was discontinued and the experiment continued under chloralose anaesthesia (70 mg.- kg-" ixv.). The animals were paralysed with gallamine triethiodide, 40 mg i v. initially and 20 mng h'1 subsequently, and artificially respired. After tracheal, venous and arterial cannulations the sural and medial plantar nerves were exposed in both hind limbs. The spinal cord was exposed at upper cervical and lumbosacral levels and covered with liquid paraffin B.Px. at 370 C. The sensorimotor cortex contralateral to the lumbo-sacral recording site was exposed and covered with 50 % liquid paraffin B.P. 50 % petroleum jelly B.P. at 370 C. An enlarged photograph of the cortex was available during the recording session and was used to record the entry points of the stimulating micro-electrode into the cortex. A bilateral pneumothorax was performed. Rectal temperature was maintained close to 380 C with a thermostatically controlled electrical blanket on the ventral body surface. Arterial blood pressure was monitored and the end-tidal CO2 maintained at 3-5-4-0 % by adjusting the stroke volume of the respiratory pump. The
CORTICAL INHIBITION OF THE SOT
3
pump volume was adjusted so that the pupils remained constricted and this occurred within the above end-tidal C02 ranges. Extracellular recordings were made from SCT cells using glass micro-pipette electrodes filled either with 4 M-NaCl (resistance 2-5 Mil) or with 5 % Procion Yellow M4RS in distilled water (resistance about 10 Mil). Methods for identifying the SCT have been described previously and included antidromic activation from the dorsolateral funiculus at C 3, with collision of ortho- and antidromic impulses, and absence of antidromic activation from C 1. Responses were evoked in SCT cells by electrical stimulation of ipsilateral cutaneous nerves (0-2 ms single shocks) or by electrical stimulation of cutaneous receptive fields through a pair of tungsten wire electrodes inserted into the skin. The peripheral stimuli were adjusted until the SCT cell under study responded with 4-20 impulses. Units were discarded if the variability in the number of impulses was too great. Once an SCT cell had been isolated and its receptive field characterized, intracortical cathodal microstimulation was performed using tungsten in glass microelectrodes (Stoney et al. 1968) with exposed tungsten tips of 15-30 /stm. The anode was a silver wire buried in the temporal muscle. Stimulating current pulses were measured as the voltage drop across a 1 kQl series resistor between the isolated stimulator and the silver anode. After preliminary trials the cortical stimulus was fixed at 3 shocks, 0-2 ms duration at 400 Hz. The conditioning-testing interval was fixed at 40 ms which is at about the time of maximal inhibitory action elicited by stimulation of the cortical surface (Brown & Short, 1974). Two series of experiments were performed. In the first series the stimulating micro-electrode was driven through the cortex in steps of 200 Fzm until inhibitory effects were seen and then in 100 /um steps through inhibitory regions using cortical stimuli of 50 or 25 VIA strength. Once inhibition was observed the cortical shocks were reduced to 10 and 5 /%A in attempts to determine the lowest strengths which would affect transmission through the SCT. Several (up to six) tracks were made in this series, usually all close to each other (500 Fm) in the transverse and sagittal directions. In the second series as much of the sensorimotor cortex as possible behind the position of the cruciate sulcus at the surface was searched with a set of electrode tracks 1-0-1-5 mm apart in the sagittal and coronal directions in attempts to surround all inhibitory areas with negative tracks. For these experiments the stimulating current was fixed at 25 /zA and the stimulation sites were every 250 Fm in depth. In both series of experiments, for each stimulation site ten unconditioned and ten conditioned responses were recorded. Initially the ten control responses were recorded before the ten conditioned responses. In the last four experiments, all in the second series, the control and conditioned responses were alternated. The recorded impulses from the SCT cell were converted to standard pulses which were counted on a counter-timer. The number of impulses in unconditioned and conditioned responses were compared using Student's t test (grouped or paired t tests where appropriate; two-tailed tests, P < 0 05 accepted as significant; the variance of the control and conditioned responses were assumed equal). Responses were also recorded on tape for subsequent analysis. After the completion of the last stimulation track, in the second series of experiments, electrolytic lesions were made by passing 10 /FA for 10 a (electrode negative) at known distances apart in the last track. In all experiments the stimulating electrode was left at its deepest position in the last track. The brain was perfused with normal saline followed by 10 % formol-saline. The next day the block of brain containing the electrode tracks was removed, the rostral and caudal surfaces of the block being cut in the coronal plane of the electrode tracks. After further
4
A. G. BROWN AND OTHERS
fixation in formol-saline four of the first series of brains were cut at 25 jsm (frozen sections) and four of the first series and all the second series were cut at 15 jam after paraffin wax embedding. All sections were cut in the coronal plane and stained with Cresyl Violet. The stimulating tracks were reconstructed. Cytoarchitectonic areas of the sensorimotor cortex were identified in each individual brain by more than one of us, and it should be noted that the criteria of Hassler & Muhs-Clement (1964) were used, not their published illustrations. We found their descriptions to be accurate. Nomenclature. In this paper the term area means a cytoarchitectonic area. Site means the position of the intracortical electrode tip during microstimulation. An inhibitory 8ite is a site from which statistically significant inhibition of an SCT cell was elicited. A negative 8ite is a site from which no significant inhibition was elicited. An inhibitory region is a group of two or more inhibitory sites adjacent in the vertical or horizontal direction without intervening negative sites. B
A
C
50
~40 r-30 0 20 C
10
0
5 10
25 4uA
50
10 i
25
uA
50
5 10
25
4uA
Text-fig. 1. The relationship between the amount of inhibition of SCT units and strength of intracortical microstimulation current. A shows results when three different current strengths produced significant inhibition, B and C show results when 2 out of 3 current strengths produced significant inhibition. The relationships are non-linear with a reduction in slope as current is increased. Each curve represents data obtained from a single SCT unit by stimulation of a single cortical site. The sixteen curves are from sixteen cortical sites and seven SCT units. Circles: significant inhibition (P < 0.05). Triangles: non-significant values. RESULTS
The aim of the first series of eight experiments was to find those regions of cortex from which inhibitory actions could be elicited at minimal current strengths and initially currents of 50 #uA were used to search for these regions. The electrode was driven through the cortex and conditioning shocks given every 200 Mum until inhibition was observed. Inhibitory effects were usually observed suddenly at a particular electrode position and went away equally abruptly as the electrode passed through the inhibitory region. When clear inhibition was obtained with 50 ,sA currents then the electrode was moved in steps of 100 Aim and currents of 25, 10
5 CORTICAL INHIBITION OF THE SOT and sometimes 5 1sA were used to determine the minimal current necessary for eliciting inhibition. The amount of inhibition (%) produced by histologically confirmed intracortical micro-stimulation, 50 pA or less, was considerably smaller than that produced by surface stimulation in earlier experiments (Brown & Short, 1974). Both 50 and 25 aA produced significant inhibition at twenty-one cortical sites. At these sites 50 pA gave an average of 32-3 % inhibition ( ± 9 7, S.D.) while 25 pA gave an average of 26 3 % inhibition (± 10-7, S.D.). At twelve cortical sites where both 25 and 10 /tA produced significant inhibition the average amounts of inhibition were 27-5 (± 9 5 %, S.D.) and 20.7 ( ± 7 0 %, S.D.), respectively. Text-fig. 1 shows percentage inhibition plotted as a function of stimulus current for all sixteen sites, obtained from 7 SCT units, where significant inhibition was elicited by three currents (Text-fig. 1 A) or by two of three currents (Text-fig. 1 B, C). It is evident that the relationship is non-linear, with a reduction in slope as current is increased. The number of sites giving rise to significant inhibition was greater with larger currents. In general 25 ,uA produced smaller inhibitory regions than 50 ,LA and in turn 10 and 5 ,uA produced even smaller ones. The positions giving maximal inhibition with the smaller currents were always similar to those giving maximal inhibition with the larger currents. Of forty-one cortical sites at which significant inhibition was produced with either 50 or 25 flA, the 50 ,jA stimulus was effective at thirty-seven (90 %) of them and the 25 pA at twenty-four (59 %). The 25 1tA stimulus was effective at four sites where 50 /tA did not produce significant inhibition. More generally, the 50 pA stimulus revealed six inhibitory regions, in four brains, which were.also revealed by the 25 ,tA stimulus. Similarly, of sixty-four sites at which significant inhibition was produced with either 25 or 10 #uA, the 25 pA stimulus was effective at fifty-nine (92 %) and the 10 pA at seventeen (25 %). There were five sites at which only the 10 #uA stimulus produced significant effects. Although eleven distinct regions were revealed using 25 #uA only six of these were revealed using 10 #uA. A further reduction of current strength to 5 #uA led to significant inhibition at only one of these six regions. Location of inhibitory regions in cortical cytoarchitectonic areas. In the first series of experiments none of the cortical inhibitory regions was entirely surrounded by negative sites. The reconstructed brains did not allow an estimate to be made of the extent and position of each inhibitory region. In mapping the cortex in three dimensions a compromise must be made between the conflicting aims of precise localization of inhibitory regions and coverage of a large volume of cortex. Experience gained in the first series suggested that a grid of tracks about 1 mm apart was the finest
A. G. BROWN AND OTHERS which would permit an extensive search within the 12 h or so for which a single SCT cell could be recorded. In the second series of seven experiments (a single SCT cell was recorded in each) the strength of shock was fixed at 25 ,#A at sites 250 ,tm apart within tracks, since this had not missed any inhibitory regions detected by larger currents or smaller steps. Using these parameters cortical sites which produced significant inhibition nearly always had at least one other similar site in one neighbouring track (see Text-figs. 2-4 and P1. 1). Only 6
2.1mm
A
c
Inhibitio
P
