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The effects of curare on cortical activity
THE EFFECTS OF CURARE ON CORTICAL ACTIVITY 1 NOEL MORLOCK AND ARTHUR A. WARD, JR., M . D .
Division of Neurosurgery, University of Washington School of Medicine, Seattle, Washington (U.S.A.) (Received for publication: May 18, 1960)
INTRODUCTION
It is becoming increasingly apparent that there are two spatially separate and functionally different types of neuronal membrane in the cerebral cortex. The membrane of certain portions of the neuron is capable of supporting all-or-none spike responses and this would appear to include portions of the soma of the cell as well as its axon. At least portions of the apical dendrite and the dendritic arborizations appear to be characterized by the production of graded responses of a non-propagating nature (Purpura and Grundfest 1956; Eccles 1951; Clare and Bishop 1955). Bishop (1956) has pointed out that this duality of function of different parts of the same nerve cell may be the result of evolutionary development in which the graded response membrane is the more primitive while the spike generating membrane represents the more recent evolutionary change to provide rapid communication between parts of the brain which have become increasingly distant from one another as the brain size enlarged. An additional major difference between these two types of membrane may be that the graded response membrane is electrically inexcitable while the spike generating membrane is electrically excitable as proposed by Grundfest (1957). Included in the supporting data are the observations of Grundfest and Purpura (1956) that intravenous curare 'in doses of 2- 3 mg/kg will reversibly block the direct cortical response (DCR) which is generally thought to represent the summed post-synaptic potentials of the apical dendrites in the cortex (Eccles 1951; Clare and Bishop 1955; Ochs 1956). If curare does, in fact,
block axo-dendritic conduction in cortex, this would lend powerful support to the concept of (1) the chemical or neural nature of axo-dendritic synaptic conduction in the cortex; (2) the electrical inexcitability of cortical dendrites; and (3) that the DCR is truly an envelope of cortical dendritic post-synaptic potentials (p.s.p.'s). However, previous observations regarding alterations of cortical function following the administration of curare are not in complete agreement with the data presented by Purpura and Grundfest (1956). Because of the discrepancies in the data and the potential importance of the conclusions to be drawn from such observations, there would appear to be a need to delineate the action of curare on cortical function in a more definitive manner. For these reasons, we have carried out three types of experiments: first, the observation of changes in the DCR produced by the intravenous administration of curare; second, the effects on the DCR of curare applied topically rather than given intravenously; and third, the modifications of firing of single cells in the cerebral cortex following the topical application of curare. METHODS
Experiments were carried out on thirty cats anesthetized with 30 mg/kg of pentobarbitone intraperitoneally. In all the animals the left calvarium and dura were removed and, if needed, the right ulnar nerve exposed. Recording of the DCR in the anterior supra-sylvian gyrus or evoked activity in the posterior cruciate gyrus usually began three to four hours subsequent to the administration of anesthesia. The DCR and the sensory evoked response a This investigation was supported, in part, by a grant B-193 (C7) from the National Institute of Neurological were recorded by means of a glass electrode having a pore 1 mm in diameter and filled with Diseases and Blindness of the National Institutes of Health, U.S. Public Health Service. saline. This electrode was counter-balanced on a 6O
CURARE AND CORTICALACTIVITY lever mechanism so as to maintain constant pressure on the cortex (Brinley, Kandel and Marshall 1958). Microelectrodes used to record from single cortical nerve cells, extracellularly, were drawn from glass capillary tubing and filled with 3 M KCI. Tip diameters from 3 - 8 # were best suited for our experiments. To minimize cortical pulsations a curved pressure plate having a 2 mm hole in its center was lowered convexity downward onto the cortex (Phillips 1956). By means of a Pfieffer micromanipulator the microelectrodes were lowered perpendicular to the cortical surface through the hole in the pressure plate. The size of the hole left enough cortex exposed so that topical solutions could be placed directly on the cortex about the microelectrode. Both commercial solutions of curare and pure preparations of curare were used without observable difference. Recording equipment for both the gross surface and unit activity consisted of condensercoupled amplifiers feeding into a dual beam oscilloscope. Single unit activity was also monitored by an audio amplifier and speaker. Continuous blood pressure recording through a femoral artery cannula was accomplished with the use of a Statham pressure transducer connected to a Grass polygraph. To obtain the DCR, Ag-AgC1 stimulating electrodes, counterbalanced on a lever, were used. The tips of these electrodes were spaced 1-2 mm apart and the best responses were obtained when the stimulating and recording electrodes were 3 - 6 mm apart. A Grass stimulator and stimulus isolation unit were employed and pulses of 0.1-0.2 msec duration with intensities of 2 - 1 5 V were found sufficient.
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Brinley et al. (1958) have shown that rigidly held electrodes may introduce artefactual changes due to alterations in the brain-electrode contact resulting from changing brain volume. In light of this we too used counterbalanced electrode assemblies to eliminate, if possible, any mechanical artefacts that might be produced by changes in brain volume following intravenous curare. Arterial blood pressure recording in our experiments revealed a blood pressure drop in the vicinity of 30-50% within 3 - 4 minutes of the time of curare injection. Since the alterations in the D C R reported by others might be due to this
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RESULTS A. Effects o f intravenous curare on the direct cortical response These experiments were designed to evaluate the alterations, if any, in the D C R subsequent to intravenous curare injection. This proved to be a difficult task because of the various systemic effects of curare. There appears to be a release of histamine after curare injection and this results in a rapid fall in the blood pressure and secondary shrinkage of brain size (Brinley, Kandel and Marshall 1958). Under these circumstances,
A Fig. 1 This Figure shows some of the variable effects of intravenous curare on the direct cortical response. A. The first record is the control direct cortical response obtained from the suprasylvian gyrus. The next three records were taken at 4 min, 12 min, and 25 min respectively, from the time of injection of curare in 3 mg/kg dosage. B. Again the first record is the control direct cortical response obtained from the suprasylvain gyrus. At 5 and 15 min after injection of curare in dosage of 3 mg/kg the following two records were made.
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N. MORLOCK AND A. A. WARD
significant drop in the blood pressure, considerable effort was expended to maintain the blood pressure by various means. Use of Dextran, trunk and limb pressure wrappings and vasopressors, however, were either found to be ineffectual in controlling blood pressure or they introduced additional unwanted variables. The DCR monitored in our experiments was obtained from the anterior supra-sylvian gyrus by means of a glass pore recording electrode spaced 3 - 6 mm away from the bipolar stimulating electrode. We found that intravenous curare produced varying effects on the DCR, ranging from no alteration to better than 50~o attenuation of the DCR. Shown in Fig. 1 are the observations made in two representative experiments in this series of twelve. The responses in Fig. 1B show minimal attenuation of the DCR within 30 minutes of the time of curare injection. In contrast to this, the experiment illustrated in Fig. 1A shows a 57~/o ~ttenuation of the DCR 4 minutes following injcction of 3 mg/kg curare with near return to normal by 25 minutes. The non-uniformity of these changes in the DCR as observed in our experiments might be explained by the fact that each animal reacts systemically in a somewhat different manner to the blood pressure drop so induced. That this might be the case is indicated by our failure to show any consistent correlation between the degree of blood pressure drop and changes in the DCR. On the basis of these preliminary observations, we were unable to convince ourselves that alterations of blood pressure were the sole variable in producing the inconstant changes in amplitude of the DCR. Since we have been unable to obtain reproducible and consistent alterations in the DCR following intravenous curare in these dosage levels, we feel that interpretations from such data must be viewed with caution.
Here again, as in the previous experimental series, balanced electrodes were used to obtain the negative dendritic response following nearby stimulation of the suprasylvian gyrus. The distance between the stimulating and recording electrodes was 3 - 6 mm and the bipolar stimulating electrode tips were about 1 mm apart. Curare was topically applied by means of small filter paper pledgers saturated with the drug and placed so as to surround the recording electrode. Because of low pH and preservatives in the commercial preparations of curare, pure bicarbonate buffered solutions of curare were tried with no observable differences. As a result of this, commercial preparations of 3 mg/cc concentration were used. In this series of experiments, reproducibility was very good and one is impressed with consistent data in contrast to that obtained after the intravenous use of curare. Topical curare was followed by enlargement of the negative dendritic response in every case. This augmentation A
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B. Topical curare and its effects on the direct cortical response
Because of our inability to obtain definitive information concerning the effects of intravenous curare on cortical function, experiments were carried out utilizing topical curare. In this manner the uncontrollable systemic side effects of curare were circumvented.
Fig. 2 This figure shows the augmenting affect of curare applied topically to the cortex on the direct cortical response. Record A, control response; B, 3 min after application; C, 4 rain; D, 7 rain; E, 10 min.
CURARE AND CORTICAL ACTIVITY
63
of the D C R was observed to be in the range of three to four times that seen in the control. Representative results from this series of ten cats are shown in Fig. 2. Augmentation of the D C R is evident within five minutes of the application of the curare-soaked pledgets and the maximum effect is seen by 10 minutes. A concomitant augmentation was also observed in the initial positive deflection of the DCR, but not to the degree seen with the negative component. The development of a second positive deflection in the D C R as shown in Fig. 2D was an additional common finding in our experiments. Infrequently, as shown in Fig. 2E, a second negative wave would appear, the significance of which is not known. The alterations produced by curare in the D C R would remain for several hours after the curare pledgets were removed and the cortex washed with saline. This time interval was too great to confidently record the return of the D C R to its original proportions.
the left somatosensory cortex of the cat until a neuron driven by the peripheral stimulation was found. Units selected for use in these experiments were those that followed the peripheral stimulus in a one to one relationship with responses consisting of 1 - 3 spike potentials. Also only those units firing at or near the negative peak of gross evoked potential were used. Before curare was placed upon the cortex, the unit was followed for at least 5 min so as to determine whether there was a constant pattern of firing of the u~.t in response to the peripheral simulus. Once the stability was felt to be adequate, curare was placed in the concave disc. Our findings revealed that curare caused an increase in the number of spikes evoked by the peripheral stimulus. Fig. 3 illustrates the typical result noted after curare was applied topically. Usually within 4 - 6 min the unit began to increase the number of discharges in its response to peripheral nerve stimulation and in 10 min was firing in groups of 10 or more spike potentials.
C. Modifications of single unit firing in the cerebral cortex after topical application of curare To gain more specific information regarding the effect of curare on the function of the cerebral cortex it was decided to study single unit behavior after topical application of curare. In this series eight animals were used and a total of eleven experiments performed. In order to achieve the mechanical stability required to record from a single unit extracellularly for upwards of 30 min a concave disc having a small hole in its center was lowered convexity downward upon the cortex. This method was found quite adequate in eliminating disturbances resulting from cortical pulsations. The microelectrodes were lowered through the hole in the disc and down into the cortex. Single cortical units were held as long as 4½ hours. The duration of the unit potentials recorded was about 0.8-1.0 msec and their spike amplitude varied from 0.3-1 mV with our equipment. The cortex immediately surrounding the microelectrode could be bathed in curare solution by placing a few drops in the disc which had its concavity facing away from the brain surface. The right ulnar nerve was stimulated at 1 sec intervals and the microelectrode was probed into
Fig. 3 This figure illustrates the effects of topically applied curare on spiking activity of single cortical units. Record A, control response; B, 3 rnin after application; C, 5 rnin; D. 8 rain, E, 10 rain; F, 12 min.
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N. MORLOCK AND A. A. WARD
The effects of curare lasted for several hours and no units were held well enough to feel secure in making any statements about the return of the unit to its original frequency of response. Here again in this series as in the series involving topical curare and gross surface recording, reproducibility was very good in contrast to experiments in which intravenous curare was used. DISCUSSION
The employment of pharmacologic substances in neurophysiology for the elucidation of some of the complex functions present in the nervous system has been of great significance in the past and no doubt will continue to be in the future. Caution is necessary, however, in making deductions regarding their mechanism of action as many pitfalls exist. In our three sets of experiments it very soon became apparent that the topical application of curare to the specific neural tissue of interest gave considerably more reliable and reproducible data than when given intravenously. This same impression is obtained when the literature dealing with the effect of curare on central nervous system function is reviewed. One is immediately struck by the discrepancies existing in the data of the many investigators who have worked with this drug. The data obtained by various workers using curare intravenously appears to fall into three major divisions. There are many investigators who found no alterations in the central nervous system activity subsequent to the injection of curare in doses ranging from that just able to paralyze the animal to massive amounts several times the paralyzing dose (Smith et aL 1947; Wesco et al. 1948; Brinley et al. 1958; Naess 1950; Girden 1948). It should be added that two of these groups of workers used various means to counteract the blood pressure drop and poor oxygenation resulting from parenteral administration of curare, feeling that these systemic alterations may well be the cause of any observed alterations of electrical activity. Despite this host of data compiled to show the lack of influence on central nervous system function induced by I.V. curare, other investigators report alterations in brain function following the administration of this drug. Among those workers who observed changes in central
nervous system activity, some have felt that curare is a central depressant (McIntyre et ak 1946; McCawley 1949; Fan and Feng 1957; Ostow and Garcia 1949; Purpura and Grundfest 1956; Gellhorn 1949; Pick and Unna 1945) while others describe the action of curare as a central excitant (Eccles 1946; Cohnberg 1946; Tillie 1890; McCawley 1949). The reasons for the disparity in the results reported by these workers are not clear. Three explanations have been entertained but each, unfortunately, is based upon tenuous evidence. Probably the most common of the three explanations is that the varying degrees of anoxia and hypotension which follow the intravenous administration of large doses of curare account for the variations in the DCR. However, we were unable to correlate consistently the amplitude of the DCR with changes in blood pressure induced by intravenous curare. This is somewhat at variance with the recent report of Ochs (1959) who reports that a reduction of blood pressure to levels of 50-60 mm Hg following intravenous curare was associated with decreases in amplitude of the DCR; while the DCR is abolished if the blood pressure is reduced below 40 mm Hg. However, they also report that a reduction of blood pressure to levels of 35-40 mm Hg by bleeding usually produces an enhancement of the DCR. Nevertheless, he postulates that the decrease in DCR following the intravenous injection of curare is secondary to the reduction of blood pressure. On the other hand, Ostow and Garcia (1949), Purpura and Grundfest (1956) and Brinley et al. (1958) felt that the DCR is quite resistant to the hypoxia and hypotension which may be encountered following intravenous curare. The second explanation is that the bloodbrain barrier is relatively impermeable to curare and variable amounts of the drug will cross into the central nervous system dependent upon the concentration of drug in the serum. However, Brinley et al. (1958) have shown that local destruction of the blood-brain barrier does not alter the effects of intravenous curare. The third and last explanation is that shrinkage of the brain occurs following intravenous curare and may alter the conditions of stimulation and recording. Brinley, Kandel and Marshall
CURARE AND CORTICAL ACTIVITY
(1958) have shown that brain shrinkage does, in fact, occur following intravenous curare and that this may change brain-electrode contact if rigidly held electrodes are utilized. In contrast, Grundfest, in his discussion of the results of Brinley et al. (1958), has reported that, in heparinized animals, an increase occurs in the DCR despite a large drop in the blood pressure. He points out that the hypotension in such animals, if it produced brain shrinkage with concomitant alteration of brain-electrode contact, should result in a decrease rather than an augmentation of the DCR if this explanation is valid. Thus, whether intravenous curare produces its CNS effects directly or indirectly via its systemic side-actions remains unresolved at present and our experiments provide no further insight into this perplexing problem. In direct contrast to the divergent results obtained with the use of intravenous curare, all of the reported observations following topical application of the drug are in unanimous agreement that curare is a central stimulant (Feldberg, Malcolm and Sherwood 1956; Chang 1953; Salama and Wright 1950; Eccles 1946; Tillie 1890; McGuigan 1916; Enomoto and AjmoneMarsan 1959; Joseph and Meltzer 1911). Our results with topical curare also are in agreement that this drug is a central stimulant. Our data are consistent with the reports of Feldberg, Malcolm and Sherwood (1956) and Chang (1953) that curare applied topically to the cortex produces a many-fold increase in the DCR. Although solutions containing high concentrations of curare were used in our experiments, Feldberg et al. (1956) have demonstrated that the augmentation of the DCR which follows topical application of curare is not a function of concentration. At concentrations which are threshold for modifying the DCR (10/tg/ml) augmentation was obtained and this effect increased with increasing concentrations of the drug. Furthermore we have observed that there is an increase in the firing of single cortical neurons following topical curare which confirms similar data reported by Enomoto and Ajmone-Marsan (1959). It would thus appear that the divergent data reported following the administration of intravenous curare does not form a consistent basis upon which significant interpretations can
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be made. In contrast, the uniformly consistent results reported following the topical application of curare would appear to provide a reliable body of data from which deductions can be made regarding the specific action of curare upon the central nervous system. Any referral to the central action of curare in the remainder of this discussion applies to the topical actions of this drug only. Whether curare blocks inhibitory synapses or excites the excitatory synapses in producing the "excitant" action observed is now a question that is close to being resolved, particularly since the techniques of single unit recording have been brought to bear upon this problem. Eccles (1946) observed that subsequent to bathing the spinal cord of the frog in curare, the duration of the synaptic potential and the discharge of impulses increased in the spinal motoneurones subjected to the influence of a single dorsal root volley. Enomoto and Ajmone-Marsan (1959) and our work has shown that the rate of discharge of cortical neurons also increases after local application of curare. Assuming that these cortical neurons function not unlike spinal neurons (Eccles 1951), one might assume that an enlargement of the synaptic potential of these cortical neurons occurs as does that of the spinal motoneurone when bathed with curare as demonstrated by Eccles (1946). This would be consistent with similar investigations with strychnine. Strychnine has been shown by Eccles, Fatt and Koketsu (1954) to decrease the IPSP of single motoneurones in the spinal cord, despite the actual increase in the discharge rate of the Renshaw cells, known to have an inhibitory influence on these motoneurones. They then concluded that strychnine specifically blocked the inhibitory" synapse on the motoneurone. Cortical neurones when recorded intracellularly after strychnine applicatioff, as noted by Li (1959), increase their discharge rate and the prepotential prior to the discharge also increases. This prepotential is no doubt analogous to the p.s.p, recorded in spinal motoneurones by Eccles et aL (1954) and one can only assume, if strychnine blocks inhibitory synapses specifically, the increased magnitude of the cortical neuron prepotential seen by Li also represents a block of inhibitory hyperpolarization normally effecting these cells.
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N. MORLOCK AND A. A. WARD
Lacking definitive intracellular investigation in either cord or cortex of the response of single cells to curare, one must infer from data accumulated thus far that curare, in the same manner as strychnine, increases the synaptic potential of nerve cells. One might thus postulate that curare blocks inhibitory synapses, thus allowing the excitatory synapses to produce the depolarizing synaptic potential without opposition and in such manner make the nerve cell more excitable. Some supporting data for this viewpoint is available. Eccles (1946) and Eccles et al. (1954) have shown that strychnine applied to the spinal cord causes an augmentation of the ventral root potential following dorsal root stimulation. Furthermore, they have shown that the synaptic potential of individual motoneurones in cord is augmented by strychnine and this augmentation is directly correlated with the enhancement of the previously described ventral root potential. In a similar manner' they have shown that curare produces an augmentation of the ventral root potential from which one can conclude that the synaptic potential of the motoneurones is also increased. However, there is no alteration of Renshaw cell activity which is normally inhibitory on the motoneurone. One can therefore conclude that curare blocks these inhibitory synapses upon the motoneurone, this reduction of hyperpolarization resulting in the increase of synaptic potential assumed to exist. Eccles et al. (1954) have presented data indicating that this is the mechanism of action of strychnine. The striking similarity of action on the central nervous system of the two related alkaloids curare and strychnine has been repeatedly reported since Tillie, in 1889, pointed out that the difference in their central action appeared to be a function of the relative dosages and the temporal order in which their various actions occurred. We have already pointed out the similarity of action of strychnine and curare on single cortical neurons. Thus, on the basis of the currently available data, we postulate that these two alkaloids alter the synaptic potential of cortical neurons by blocking inhibitory synapses. However, further data is needed since there is not yet complete agreement even with respect to the mode of action of strychnine.
SUMMARY
(1) Intravenous curare produced variable degrees of depression of the direct cortical response. From these experiments it was observed that brain shrinkage and/or blood pressure changes secondary to the intravenous injection of curare can not explain the lack of reproducible results. Since uncontrolled variables appear to be operating, it is felt that interpretations from such data must be viewed with caution. (2) Application of curare topically to the cortex, in contrast to its intravenous administration, caused consistent and reproducible augmentation of the direct cortical response. (3) Microelectrode recording demonstrated that topical curare increased the number of responses of a cortical neuron to a single peripheral stimulus. (4) From these results, the specific action of curare on the central nervous system is considered to be excitatory, possibly as the result of blockade of inhibitory synapses. REFERENCES BERNHARD, C. G. and TRAVERNER, D. The action of d-
tubocurarine on the monosynaptie extensor reflex., Brit. J. Pharraacol., 1951, 6: 540-550. BISHOP,G.H. Natural history of the nerve impulse. PhysioL Rev., 1956, 36: 376-399. BRINLEY, F.J., KANDEL, E . R . and MARSHALL, W. H. The effect of intravenous d-tubocurarine on the electrical activity of the cat cerebral cortex. Trans. Amer. neuroL Ass., 1958, 53-58. CHANG, H. Similarity in action between curare and strychnine on cortical neurons. J. NeurophysioL, 1953, 16: 221. CLARE, M. H. and BISHOP, G. H. Dendritic circuits: The properties of cortical paths involving dend rites. A mer. J. Psychiat., 1955, 3: 818-825. COHNBERG,R. E. Stimulation of the central nervous system by curare Ontocostrin). J. Lab. clin. Med., 1946, 31: 866-877. ECCLES, J.C. Synaptic potentials of rnotoneurones. J. NeurophysioL, 19416, 9: 87-120. ECCLES, J. C. Interpretation of action potentials evoked in the cerebral cortex. Electroenceph. clin. Neurophysiol., 1951, 3: 449-464. ECCLES, J. C., FAT'I', P. and KOKETSU, K. Cholinergie and inhibitory synapses in a pathway from motor-axon collaterals to motoneurones. J. Physiol., 1954, 126: 524-562. ENOMOTO, T. F. and AJMONE-MARSAN, C. Epileptic activation of single cortical neurons and their relationship with electroencephalographic discharges. Electroenceph, clin. Neurophysiol., 1959, 11 : 199-218.
CURARE AND CORTICAL ACTIVITY FAN, S. and FENO, T. Concerning conduction and electrical excitability in the terminal portion of the apical dendrites of the pyramidal neurons. Acta physiol, sin., 1957, 21 : 423-434. FELDBERG, W., MALCOLM, J.L. and SHERWOOD, S. L. Some effects of tubocurarine on the electrical activity of the cat's brain. J. Physiol., 1956, 132: 130-145. GELLHORN, E. The influence of curare on hypothalamic excitability and the electroencephalogram. Electroenceph, clin. Neurophysiol., 1958, 10: 697-703. GIRDEN, E. The electroencephalogram (EEG) in curarized animals. J. Neurophysiol., 1948, 11: 169-173. GRUNDFEST, H. Electrical inexcitability of synapses and some consequences in the central nervous system. Physiol. Rev., 1957, 37: 337-361. JOSEPH, D. R. and MELTZER, S. J. On a difference between effects of intravenous and intra-aortic injections of curarin in frogs. J. Pharmacol., 1911, 3: 465. LI, C. Cortical intracellular potentials and their responses to strychnine. J. Neurophysiol., 1959, 22: 436-450. MC CAWLEY,E. L. Central action of curare. J. Pharmacol., 1949, 97: 129. McGuIGAN, H. Central action of curare. J. Pharmacol., 1916, 8: 471-477. MCINTYRE, A. R., DUNN, A. L., and TULLAR, P. E. The effect of d-tubocurarine on the electrical activity of dog's brains. Fed. Proc., 1946, 5: 67. NAESS, K. The effect of d-tubocurarine on the mono- and polysynaptic reflex of the spinal cord including a comparison with the effect of strychnine. Acta physiol, scand., 1950, 21: 34-40.
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OcI-m, S. The direct cortical response. J. Neurophysiol., 1956, 19: 513-523. OCHS, S. Curare and low blood pressure effects on direct cortical responses, Amer. J. Physiol., 1959, 197: 1136-1140. OSTOW, M. and GARCIA, F. Effect of curare on cortical responses evoked by afferent stimulation. J. Neurophysiol., 1949, 12: 225-229. PHILLIPS, C. G. Cortical motor threshold and the thresholds and distribution of excited Betz cells in the cat. Quart. J. exp. Physiol., 1956, 40: 70-84. PICK, E. P. and Ur,~A, K. The effect of curare and curarelike substances on the central nervous system, d~ Pharmacol., 195, 83: 59-70. PURPURA, D. P. and GRUNDFEST, H. Nature of dendritic potentials and synaptic mechanisms in cerebral cortex of cat. J. Neurophysiol., 1956, 19: 573-593. SALAMA,S. and WRIOnT, S. Action of d-tubocurarine chloride on the central nervous system of the cat. Brit. J. Pharmacol., 1950, 5: 49-61. S~arrH, S. M., BROWN, H. O., TOMAN, J. E. P., and GOODMAN, L. The lack of cerebral effects of d-tubocurarine. Anesthesiology, 1947, 8: 1-14. TmLm, J. A. Contribution to the pharmacology of curare and its alkaloids. J. Anat. Physiol., 1890, 24: 379-406. W ~ c o , C.W., GREEN, R.E., MCNAMARA, B.P. and KROP, S. The influence of atropine and scopolamine on the central effect of DFP. J. Pharmacol., 1948, 92: 63-72.
Reference: MORLOCK, N. and WARD, A. A. The effects of curare on cortical activity. Electroenceph. clin. Neurophysiol., 1961, 13: 60-67.
Division of Neurosurgery, University of Washington School of Medicine, Seattle, Washington (U.S.A.) (Received for publication: May 18, 1960)
INTRODUCTION
It is becoming increasingly apparent that there are two spatially separate and functionally different types of neuronal membrane in the cerebral cortex. The membrane of certain portions of the neuron is capable of supporting all-or-none spike responses and this would appear to include portions of the soma of the cell as well as its axon. At least portions of the apical dendrite and the dendritic arborizations appear to be characterized by the production of graded responses of a non-propagating nature (Purpura and Grundfest 1956; Eccles 1951; Clare and Bishop 1955). Bishop (1956) has pointed out that this duality of function of different parts of the same nerve cell may be the result of evolutionary development in which the graded response membrane is the more primitive while the spike generating membrane represents the more recent evolutionary change to provide rapid communication between parts of the brain which have become increasingly distant from one another as the brain size enlarged. An additional major difference between these two types of membrane may be that the graded response membrane is electrically inexcitable while the spike generating membrane is electrically excitable as proposed by Grundfest (1957). Included in the supporting data are the observations of Grundfest and Purpura (1956) that intravenous curare 'in doses of 2- 3 mg/kg will reversibly block the direct cortical response (DCR) which is generally thought to represent the summed post-synaptic potentials of the apical dendrites in the cortex (Eccles 1951; Clare and Bishop 1955; Ochs 1956). If curare does, in fact,
block axo-dendritic conduction in cortex, this would lend powerful support to the concept of (1) the chemical or neural nature of axo-dendritic synaptic conduction in the cortex; (2) the electrical inexcitability of cortical dendrites; and (3) that the DCR is truly an envelope of cortical dendritic post-synaptic potentials (p.s.p.'s). However, previous observations regarding alterations of cortical function following the administration of curare are not in complete agreement with the data presented by Purpura and Grundfest (1956). Because of the discrepancies in the data and the potential importance of the conclusions to be drawn from such observations, there would appear to be a need to delineate the action of curare on cortical function in a more definitive manner. For these reasons, we have carried out three types of experiments: first, the observation of changes in the DCR produced by the intravenous administration of curare; second, the effects on the DCR of curare applied topically rather than given intravenously; and third, the modifications of firing of single cells in the cerebral cortex following the topical application of curare. METHODS
Experiments were carried out on thirty cats anesthetized with 30 mg/kg of pentobarbitone intraperitoneally. In all the animals the left calvarium and dura were removed and, if needed, the right ulnar nerve exposed. Recording of the DCR in the anterior supra-sylvian gyrus or evoked activity in the posterior cruciate gyrus usually began three to four hours subsequent to the administration of anesthesia. The DCR and the sensory evoked response a This investigation was supported, in part, by a grant B-193 (C7) from the National Institute of Neurological were recorded by means of a glass electrode having a pore 1 mm in diameter and filled with Diseases and Blindness of the National Institutes of Health, U.S. Public Health Service. saline. This electrode was counter-balanced on a 6O
CURARE AND CORTICALACTIVITY lever mechanism so as to maintain constant pressure on the cortex (Brinley, Kandel and Marshall 1958). Microelectrodes used to record from single cortical nerve cells, extracellularly, were drawn from glass capillary tubing and filled with 3 M KCI. Tip diameters from 3 - 8 # were best suited for our experiments. To minimize cortical pulsations a curved pressure plate having a 2 mm hole in its center was lowered convexity downward onto the cortex (Phillips 1956). By means of a Pfieffer micromanipulator the microelectrodes were lowered perpendicular to the cortical surface through the hole in the pressure plate. The size of the hole left enough cortex exposed so that topical solutions could be placed directly on the cortex about the microelectrode. Both commercial solutions of curare and pure preparations of curare were used without observable difference. Recording equipment for both the gross surface and unit activity consisted of condensercoupled amplifiers feeding into a dual beam oscilloscope. Single unit activity was also monitored by an audio amplifier and speaker. Continuous blood pressure recording through a femoral artery cannula was accomplished with the use of a Statham pressure transducer connected to a Grass polygraph. To obtain the DCR, Ag-AgC1 stimulating electrodes, counterbalanced on a lever, were used. The tips of these electrodes were spaced 1-2 mm apart and the best responses were obtained when the stimulating and recording electrodes were 3 - 6 mm apart. A Grass stimulator and stimulus isolation unit were employed and pulses of 0.1-0.2 msec duration with intensities of 2 - 1 5 V were found sufficient.
61
Brinley et al. (1958) have shown that rigidly held electrodes may introduce artefactual changes due to alterations in the brain-electrode contact resulting from changing brain volume. In light of this we too used counterbalanced electrode assemblies to eliminate, if possible, any mechanical artefacts that might be produced by changes in brain volume following intravenous curare. Arterial blood pressure recording in our experiments revealed a blood pressure drop in the vicinity of 30-50% within 3 - 4 minutes of the time of curare injection. Since the alterations in the D C R reported by others might be due to this
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RESULTS A. Effects o f intravenous curare on the direct cortical response These experiments were designed to evaluate the alterations, if any, in the D C R subsequent to intravenous curare injection. This proved to be a difficult task because of the various systemic effects of curare. There appears to be a release of histamine after curare injection and this results in a rapid fall in the blood pressure and secondary shrinkage of brain size (Brinley, Kandel and Marshall 1958). Under these circumstances,
A Fig. 1 This Figure shows some of the variable effects of intravenous curare on the direct cortical response. A. The first record is the control direct cortical response obtained from the suprasylvian gyrus. The next three records were taken at 4 min, 12 min, and 25 min respectively, from the time of injection of curare in 3 mg/kg dosage. B. Again the first record is the control direct cortical response obtained from the suprasylvain gyrus. At 5 and 15 min after injection of curare in dosage of 3 mg/kg the following two records were made.
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N. MORLOCK AND A. A. WARD
significant drop in the blood pressure, considerable effort was expended to maintain the blood pressure by various means. Use of Dextran, trunk and limb pressure wrappings and vasopressors, however, were either found to be ineffectual in controlling blood pressure or they introduced additional unwanted variables. The DCR monitored in our experiments was obtained from the anterior supra-sylvian gyrus by means of a glass pore recording electrode spaced 3 - 6 mm away from the bipolar stimulating electrode. We found that intravenous curare produced varying effects on the DCR, ranging from no alteration to better than 50~o attenuation of the DCR. Shown in Fig. 1 are the observations made in two representative experiments in this series of twelve. The responses in Fig. 1B show minimal attenuation of the DCR within 30 minutes of the time of curare injection. In contrast to this, the experiment illustrated in Fig. 1A shows a 57~/o ~ttenuation of the DCR 4 minutes following injcction of 3 mg/kg curare with near return to normal by 25 minutes. The non-uniformity of these changes in the DCR as observed in our experiments might be explained by the fact that each animal reacts systemically in a somewhat different manner to the blood pressure drop so induced. That this might be the case is indicated by our failure to show any consistent correlation between the degree of blood pressure drop and changes in the DCR. On the basis of these preliminary observations, we were unable to convince ourselves that alterations of blood pressure were the sole variable in producing the inconstant changes in amplitude of the DCR. Since we have been unable to obtain reproducible and consistent alterations in the DCR following intravenous curare in these dosage levels, we feel that interpretations from such data must be viewed with caution.
Here again, as in the previous experimental series, balanced electrodes were used to obtain the negative dendritic response following nearby stimulation of the suprasylvian gyrus. The distance between the stimulating and recording electrodes was 3 - 6 mm and the bipolar stimulating electrode tips were about 1 mm apart. Curare was topically applied by means of small filter paper pledgers saturated with the drug and placed so as to surround the recording electrode. Because of low pH and preservatives in the commercial preparations of curare, pure bicarbonate buffered solutions of curare were tried with no observable differences. As a result of this, commercial preparations of 3 mg/cc concentration were used. In this series of experiments, reproducibility was very good and one is impressed with consistent data in contrast to that obtained after the intravenous use of curare. Topical curare was followed by enlargement of the negative dendritic response in every case. This augmentation A
~i)
B
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B. Topical curare and its effects on the direct cortical response
Because of our inability to obtain definitive information concerning the effects of intravenous curare on cortical function, experiments were carried out utilizing topical curare. In this manner the uncontrollable systemic side effects of curare were circumvented.
Fig. 2 This figure shows the augmenting affect of curare applied topically to the cortex on the direct cortical response. Record A, control response; B, 3 min after application; C, 4 rain; D, 7 rain; E, 10 min.
CURARE AND CORTICAL ACTIVITY
63
of the D C R was observed to be in the range of three to four times that seen in the control. Representative results from this series of ten cats are shown in Fig. 2. Augmentation of the D C R is evident within five minutes of the application of the curare-soaked pledgets and the maximum effect is seen by 10 minutes. A concomitant augmentation was also observed in the initial positive deflection of the DCR, but not to the degree seen with the negative component. The development of a second positive deflection in the D C R as shown in Fig. 2D was an additional common finding in our experiments. Infrequently, as shown in Fig. 2E, a second negative wave would appear, the significance of which is not known. The alterations produced by curare in the D C R would remain for several hours after the curare pledgets were removed and the cortex washed with saline. This time interval was too great to confidently record the return of the D C R to its original proportions.
the left somatosensory cortex of the cat until a neuron driven by the peripheral stimulation was found. Units selected for use in these experiments were those that followed the peripheral stimulus in a one to one relationship with responses consisting of 1 - 3 spike potentials. Also only those units firing at or near the negative peak of gross evoked potential were used. Before curare was placed upon the cortex, the unit was followed for at least 5 min so as to determine whether there was a constant pattern of firing of the u~.t in response to the peripheral simulus. Once the stability was felt to be adequate, curare was placed in the concave disc. Our findings revealed that curare caused an increase in the number of spikes evoked by the peripheral stimulus. Fig. 3 illustrates the typical result noted after curare was applied topically. Usually within 4 - 6 min the unit began to increase the number of discharges in its response to peripheral nerve stimulation and in 10 min was firing in groups of 10 or more spike potentials.
C. Modifications of single unit firing in the cerebral cortex after topical application of curare To gain more specific information regarding the effect of curare on the function of the cerebral cortex it was decided to study single unit behavior after topical application of curare. In this series eight animals were used and a total of eleven experiments performed. In order to achieve the mechanical stability required to record from a single unit extracellularly for upwards of 30 min a concave disc having a small hole in its center was lowered convexity downward upon the cortex. This method was found quite adequate in eliminating disturbances resulting from cortical pulsations. The microelectrodes were lowered through the hole in the disc and down into the cortex. Single cortical units were held as long as 4½ hours. The duration of the unit potentials recorded was about 0.8-1.0 msec and their spike amplitude varied from 0.3-1 mV with our equipment. The cortex immediately surrounding the microelectrode could be bathed in curare solution by placing a few drops in the disc which had its concavity facing away from the brain surface. The right ulnar nerve was stimulated at 1 sec intervals and the microelectrode was probed into
Fig. 3 This figure illustrates the effects of topically applied curare on spiking activity of single cortical units. Record A, control response; B, 3 rnin after application; C, 5 rnin; D. 8 rain, E, 10 rain; F, 12 min.
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N. MORLOCK AND A. A. WARD
The effects of curare lasted for several hours and no units were held well enough to feel secure in making any statements about the return of the unit to its original frequency of response. Here again in this series as in the series involving topical curare and gross surface recording, reproducibility was very good in contrast to experiments in which intravenous curare was used. DISCUSSION
The employment of pharmacologic substances in neurophysiology for the elucidation of some of the complex functions present in the nervous system has been of great significance in the past and no doubt will continue to be in the future. Caution is necessary, however, in making deductions regarding their mechanism of action as many pitfalls exist. In our three sets of experiments it very soon became apparent that the topical application of curare to the specific neural tissue of interest gave considerably more reliable and reproducible data than when given intravenously. This same impression is obtained when the literature dealing with the effect of curare on central nervous system function is reviewed. One is immediately struck by the discrepancies existing in the data of the many investigators who have worked with this drug. The data obtained by various workers using curare intravenously appears to fall into three major divisions. There are many investigators who found no alterations in the central nervous system activity subsequent to the injection of curare in doses ranging from that just able to paralyze the animal to massive amounts several times the paralyzing dose (Smith et aL 1947; Wesco et al. 1948; Brinley et al. 1958; Naess 1950; Girden 1948). It should be added that two of these groups of workers used various means to counteract the blood pressure drop and poor oxygenation resulting from parenteral administration of curare, feeling that these systemic alterations may well be the cause of any observed alterations of electrical activity. Despite this host of data compiled to show the lack of influence on central nervous system function induced by I.V. curare, other investigators report alterations in brain function following the administration of this drug. Among those workers who observed changes in central
nervous system activity, some have felt that curare is a central depressant (McIntyre et ak 1946; McCawley 1949; Fan and Feng 1957; Ostow and Garcia 1949; Purpura and Grundfest 1956; Gellhorn 1949; Pick and Unna 1945) while others describe the action of curare as a central excitant (Eccles 1946; Cohnberg 1946; Tillie 1890; McCawley 1949). The reasons for the disparity in the results reported by these workers are not clear. Three explanations have been entertained but each, unfortunately, is based upon tenuous evidence. Probably the most common of the three explanations is that the varying degrees of anoxia and hypotension which follow the intravenous administration of large doses of curare account for the variations in the DCR. However, we were unable to correlate consistently the amplitude of the DCR with changes in blood pressure induced by intravenous curare. This is somewhat at variance with the recent report of Ochs (1959) who reports that a reduction of blood pressure to levels of 50-60 mm Hg following intravenous curare was associated with decreases in amplitude of the DCR; while the DCR is abolished if the blood pressure is reduced below 40 mm Hg. However, they also report that a reduction of blood pressure to levels of 35-40 mm Hg by bleeding usually produces an enhancement of the DCR. Nevertheless, he postulates that the decrease in DCR following the intravenous injection of curare is secondary to the reduction of blood pressure. On the other hand, Ostow and Garcia (1949), Purpura and Grundfest (1956) and Brinley et al. (1958) felt that the DCR is quite resistant to the hypoxia and hypotension which may be encountered following intravenous curare. The second explanation is that the bloodbrain barrier is relatively impermeable to curare and variable amounts of the drug will cross into the central nervous system dependent upon the concentration of drug in the serum. However, Brinley et al. (1958) have shown that local destruction of the blood-brain barrier does not alter the effects of intravenous curare. The third and last explanation is that shrinkage of the brain occurs following intravenous curare and may alter the conditions of stimulation and recording. Brinley, Kandel and Marshall
CURARE AND CORTICAL ACTIVITY
(1958) have shown that brain shrinkage does, in fact, occur following intravenous curare and that this may change brain-electrode contact if rigidly held electrodes are utilized. In contrast, Grundfest, in his discussion of the results of Brinley et al. (1958), has reported that, in heparinized animals, an increase occurs in the DCR despite a large drop in the blood pressure. He points out that the hypotension in such animals, if it produced brain shrinkage with concomitant alteration of brain-electrode contact, should result in a decrease rather than an augmentation of the DCR if this explanation is valid. Thus, whether intravenous curare produces its CNS effects directly or indirectly via its systemic side-actions remains unresolved at present and our experiments provide no further insight into this perplexing problem. In direct contrast to the divergent results obtained with the use of intravenous curare, all of the reported observations following topical application of the drug are in unanimous agreement that curare is a central stimulant (Feldberg, Malcolm and Sherwood 1956; Chang 1953; Salama and Wright 1950; Eccles 1946; Tillie 1890; McGuigan 1916; Enomoto and AjmoneMarsan 1959; Joseph and Meltzer 1911). Our results with topical curare also are in agreement that this drug is a central stimulant. Our data are consistent with the reports of Feldberg, Malcolm and Sherwood (1956) and Chang (1953) that curare applied topically to the cortex produces a many-fold increase in the DCR. Although solutions containing high concentrations of curare were used in our experiments, Feldberg et al. (1956) have demonstrated that the augmentation of the DCR which follows topical application of curare is not a function of concentration. At concentrations which are threshold for modifying the DCR (10/tg/ml) augmentation was obtained and this effect increased with increasing concentrations of the drug. Furthermore we have observed that there is an increase in the firing of single cortical neurons following topical curare which confirms similar data reported by Enomoto and Ajmone-Marsan (1959). It would thus appear that the divergent data reported following the administration of intravenous curare does not form a consistent basis upon which significant interpretations can
65
be made. In contrast, the uniformly consistent results reported following the topical application of curare would appear to provide a reliable body of data from which deductions can be made regarding the specific action of curare upon the central nervous system. Any referral to the central action of curare in the remainder of this discussion applies to the topical actions of this drug only. Whether curare blocks inhibitory synapses or excites the excitatory synapses in producing the "excitant" action observed is now a question that is close to being resolved, particularly since the techniques of single unit recording have been brought to bear upon this problem. Eccles (1946) observed that subsequent to bathing the spinal cord of the frog in curare, the duration of the synaptic potential and the discharge of impulses increased in the spinal motoneurones subjected to the influence of a single dorsal root volley. Enomoto and Ajmone-Marsan (1959) and our work has shown that the rate of discharge of cortical neurons also increases after local application of curare. Assuming that these cortical neurons function not unlike spinal neurons (Eccles 1951), one might assume that an enlargement of the synaptic potential of these cortical neurons occurs as does that of the spinal motoneurone when bathed with curare as demonstrated by Eccles (1946). This would be consistent with similar investigations with strychnine. Strychnine has been shown by Eccles, Fatt and Koketsu (1954) to decrease the IPSP of single motoneurones in the spinal cord, despite the actual increase in the discharge rate of the Renshaw cells, known to have an inhibitory influence on these motoneurones. They then concluded that strychnine specifically blocked the inhibitory" synapse on the motoneurone. Cortical neurones when recorded intracellularly after strychnine applicatioff, as noted by Li (1959), increase their discharge rate and the prepotential prior to the discharge also increases. This prepotential is no doubt analogous to the p.s.p, recorded in spinal motoneurones by Eccles et aL (1954) and one can only assume, if strychnine blocks inhibitory synapses specifically, the increased magnitude of the cortical neuron prepotential seen by Li also represents a block of inhibitory hyperpolarization normally effecting these cells.
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N. MORLOCK AND A. A. WARD
Lacking definitive intracellular investigation in either cord or cortex of the response of single cells to curare, one must infer from data accumulated thus far that curare, in the same manner as strychnine, increases the synaptic potential of nerve cells. One might thus postulate that curare blocks inhibitory synapses, thus allowing the excitatory synapses to produce the depolarizing synaptic potential without opposition and in such manner make the nerve cell more excitable. Some supporting data for this viewpoint is available. Eccles (1946) and Eccles et al. (1954) have shown that strychnine applied to the spinal cord causes an augmentation of the ventral root potential following dorsal root stimulation. Furthermore, they have shown that the synaptic potential of individual motoneurones in cord is augmented by strychnine and this augmentation is directly correlated with the enhancement of the previously described ventral root potential. In a similar manner' they have shown that curare produces an augmentation of the ventral root potential from which one can conclude that the synaptic potential of the motoneurones is also increased. However, there is no alteration of Renshaw cell activity which is normally inhibitory on the motoneurone. One can therefore conclude that curare blocks these inhibitory synapses upon the motoneurone, this reduction of hyperpolarization resulting in the increase of synaptic potential assumed to exist. Eccles et al. (1954) have presented data indicating that this is the mechanism of action of strychnine. The striking similarity of action on the central nervous system of the two related alkaloids curare and strychnine has been repeatedly reported since Tillie, in 1889, pointed out that the difference in their central action appeared to be a function of the relative dosages and the temporal order in which their various actions occurred. We have already pointed out the similarity of action of strychnine and curare on single cortical neurons. Thus, on the basis of the currently available data, we postulate that these two alkaloids alter the synaptic potential of cortical neurons by blocking inhibitory synapses. However, further data is needed since there is not yet complete agreement even with respect to the mode of action of strychnine.
SUMMARY
(1) Intravenous curare produced variable degrees of depression of the direct cortical response. From these experiments it was observed that brain shrinkage and/or blood pressure changes secondary to the intravenous injection of curare can not explain the lack of reproducible results. Since uncontrolled variables appear to be operating, it is felt that interpretations from such data must be viewed with caution. (2) Application of curare topically to the cortex, in contrast to its intravenous administration, caused consistent and reproducible augmentation of the direct cortical response. (3) Microelectrode recording demonstrated that topical curare increased the number of responses of a cortical neuron to a single peripheral stimulus. (4) From these results, the specific action of curare on the central nervous system is considered to be excitatory, possibly as the result of blockade of inhibitory synapses. REFERENCES BERNHARD, C. G. and TRAVERNER, D. The action of d-
tubocurarine on the monosynaptie extensor reflex., Brit. J. Pharraacol., 1951, 6: 540-550. BISHOP,G.H. Natural history of the nerve impulse. PhysioL Rev., 1956, 36: 376-399. BRINLEY, F.J., KANDEL, E . R . and MARSHALL, W. H. The effect of intravenous d-tubocurarine on the electrical activity of the cat cerebral cortex. Trans. Amer. neuroL Ass., 1958, 53-58. CHANG, H. Similarity in action between curare and strychnine on cortical neurons. J. NeurophysioL, 1953, 16: 221. CLARE, M. H. and BISHOP, G. H. Dendritic circuits: The properties of cortical paths involving dend rites. A mer. J. Psychiat., 1955, 3: 818-825. COHNBERG,R. E. Stimulation of the central nervous system by curare Ontocostrin). J. Lab. clin. Med., 1946, 31: 866-877. ECCLES, J.C. Synaptic potentials of rnotoneurones. J. NeurophysioL, 19416, 9: 87-120. ECCLES, J. C. Interpretation of action potentials evoked in the cerebral cortex. Electroenceph. clin. Neurophysiol., 1951, 3: 449-464. ECCLES, J. C., FAT'I', P. and KOKETSU, K. Cholinergie and inhibitory synapses in a pathway from motor-axon collaterals to motoneurones. J. Physiol., 1954, 126: 524-562. ENOMOTO, T. F. and AJMONE-MARSAN, C. Epileptic activation of single cortical neurons and their relationship with electroencephalographic discharges. Electroenceph, clin. Neurophysiol., 1959, 11 : 199-218.
CURARE AND CORTICAL ACTIVITY FAN, S. and FENO, T. Concerning conduction and electrical excitability in the terminal portion of the apical dendrites of the pyramidal neurons. Acta physiol, sin., 1957, 21 : 423-434. FELDBERG, W., MALCOLM, J.L. and SHERWOOD, S. L. Some effects of tubocurarine on the electrical activity of the cat's brain. J. Physiol., 1956, 132: 130-145. GELLHORN, E. The influence of curare on hypothalamic excitability and the electroencephalogram. Electroenceph, clin. Neurophysiol., 1958, 10: 697-703. GIRDEN, E. The electroencephalogram (EEG) in curarized animals. J. Neurophysiol., 1948, 11: 169-173. GRUNDFEST, H. Electrical inexcitability of synapses and some consequences in the central nervous system. Physiol. Rev., 1957, 37: 337-361. JOSEPH, D. R. and MELTZER, S. J. On a difference between effects of intravenous and intra-aortic injections of curarin in frogs. J. Pharmacol., 1911, 3: 465. LI, C. Cortical intracellular potentials and their responses to strychnine. J. Neurophysiol., 1959, 22: 436-450. MC CAWLEY,E. L. Central action of curare. J. Pharmacol., 1949, 97: 129. McGuIGAN, H. Central action of curare. J. Pharmacol., 1916, 8: 471-477. MCINTYRE, A. R., DUNN, A. L., and TULLAR, P. E. The effect of d-tubocurarine on the electrical activity of dog's brains. Fed. Proc., 1946, 5: 67. NAESS, K. The effect of d-tubocurarine on the mono- and polysynaptic reflex of the spinal cord including a comparison with the effect of strychnine. Acta physiol, scand., 1950, 21: 34-40.
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OcI-m, S. The direct cortical response. J. Neurophysiol., 1956, 19: 513-523. OCHS, S. Curare and low blood pressure effects on direct cortical responses, Amer. J. Physiol., 1959, 197: 1136-1140. OSTOW, M. and GARCIA, F. Effect of curare on cortical responses evoked by afferent stimulation. J. Neurophysiol., 1949, 12: 225-229. PHILLIPS, C. G. Cortical motor threshold and the thresholds and distribution of excited Betz cells in the cat. Quart. J. exp. Physiol., 1956, 40: 70-84. PICK, E. P. and Ur,~A, K. The effect of curare and curarelike substances on the central nervous system, d~ Pharmacol., 195, 83: 59-70. PURPURA, D. P. and GRUNDFEST, H. Nature of dendritic potentials and synaptic mechanisms in cerebral cortex of cat. J. Neurophysiol., 1956, 19: 573-593. SALAMA,S. and WRIOnT, S. Action of d-tubocurarine chloride on the central nervous system of the cat. Brit. J. Pharmacol., 1950, 5: 49-61. S~arrH, S. M., BROWN, H. O., TOMAN, J. E. P., and GOODMAN, L. The lack of cerebral effects of d-tubocurarine. Anesthesiology, 1947, 8: 1-14. TmLm, J. A. Contribution to the pharmacology of curare and its alkaloids. J. Anat. Physiol., 1890, 24: 379-406. W ~ c o , C.W., GREEN, R.E., MCNAMARA, B.P. and KROP, S. The influence of atropine and scopolamine on the central effect of DFP. J. Pharmacol., 1948, 92: 63-72.
Reference: MORLOCK, N. and WARD, A. A. The effects of curare on cortical activity. Electroenceph. clin. Neurophysiol., 1961, 13: 60-67.