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Electrophysiological studies of neurosecretory cells in the cat hypothalamus
462
SHORT COMMUNICATIONS
Electrophysiological studies of neurosecretory cells in the cat hypothalamus The neuroendocrine cells of the hypothalamus are considered to perform a dual function in that they synthesize and secrete hormones as well as conduct nerve impulses. A number of authors have obtained electrophysiological evidence in support of this contention 3,4,7. Yagi e t al. 7 reported briefly that cells in the rat's supraoptic nucleus (SON) could be antidromicaily excited by stimulation of the posterior lobe of the pituitary. Dyball and Koizumi 4 found that neuroendocrine cells in the SON of the rat hypothalamus, which had been identified by antidromic stimulation of the posterior pituitary, were sensitive not only to osmatic changes but were also excited by stimulation of vagal afferent fibers and by injection of chemicals which increased antidiuretic hormone release. Cross e t al. 3 also studied antidromic excitation of the rat paraventricular nucleus (PVN) cells. The present report extends these studies to the cat hypothalamus. Despite previous failures to evoke antidromic excitation of the cells of the SON and PVN", 6 we finally succeeded in evoking responses from cells in these nuclei by single pulse (I msec duration) stimulation of the posterior lobe of the pituitary. The nature of these responses and interactions of the nuclei were studied in some detail. The following procedures were employed. Cats were anesthetized with chloralose (35-70 mg/kg) in most instances but occasionally with Nembutal (35 mg/kg). Fine bipolar silver or steel electrodes were imbedded in the posterior lobe of the pituitary after it had been exposed by hemispherectomy, or through the palate by removal of the sphenoid bone. Recordings were made from SON and PVN neurons by glass capillary electrodes filled with 3 M KCI or with a fast-green NaCI mixture. These electrodes were positioned stereotaxically, being introduced from the'ventral side of the hypothalamus, or visually placed from the hemispherectomized side. The precise positions of recording and stimulating electrode tips were determined histologically after each experiment. Some 34 experiments were performed and recordings obtained from 98 cells. Fig. I shows extracellular and intracellular recording of antidromic excitation of cells in the SON and PVN. These responses were evoked by single pulse stimulation of the posterior lobe of the pituitary. Latencies of the antidromic potentials varied as did the distance between stimulating and recording electrodes, but for each cell in a specific experiment latency was constant and did not fluctuate as in the case of orthodromic excitation. These latencies to antidromic stimulation fell between 10-20 msec for SON cells and 15-25 msec for PVN cells. By calculating average distances between stimulating and recording electrodes, it was estimated that conduction velocities in these neurosecretory cell axons ranged from 0.4 to 0.9 m/see. These values agree well with measurements made by others in rats and cats 3-'~.7. The antidromic potentials of the neurosecretory cells usually showed a notch in their rising phase indicative of delay in invasion of the soma by the axonal impulse. This notch became clearer when the stimulus intensity was low or when high frequency stimuli were given (Fig. 1A, C). Fig I B shows an intracellularly recorded antidromic potential. Brain Research,
20 (1970)462-466
463
SHORT COMMUNICATIONS
SON
PVN
1
mV
I mV $
i 5 msec
D
I0 msec Fig. 1. Antidromically induced action potentials recorded from the supraoptic nucleus (SON) (C and D) and paraventricular nucleus (PVN) (A and B). A and C show superimposed reactions obtained with barely threshold stimuli. Note the notch in the rising phase and that block occurred in some instances at this point. All records are from extracellular recordings except B which shows a spike induced during intracellular recording. Post-spike hyperpolarization was occasionally observed as in D.
When two successive stimuli were applied to the posterior lobe, the second response was blocked at 5-10 msec stimulus interval, the second evoking only a small response resembling an initial segment (IS) spike. This indicated that the refractory period of neurosecretory cells following antidromic excitation is of the order of 10 msec. Fig. 2 presents a series of records which show that at a critical stimulus interval the second of two antidromic responses has a longer latency (B, C, E), smaller amplitude (D, E) or is shown only as an IS spike (F). Full size uniform action potentials were evoked at stimulation rates of 50-100/sec. One unique result of this investigation was the discovery that cells of the SON and PVN interact in a rather complex fashion. It was found that some cells of the SON were excited orthodromically by pulses applied to PVN cells. Fig. 3C shows a response evoked immediately after PVN stimulation. Since recording was only from a SON cell it was the SON cell which gave the response. Interactions, however, were not invariably excitatory. It is shown in Fig. 3B, C and E that this same stimulation of the PVN inhibited antidromic excitation of the SON cell completely (B, C) or block propagation of the antidromic invasion at the initial segment (IS spike in E). Records A and D show control antidromic excitations of the SON cell under study. Brain Research, 20 (1970) 462-466
464
SHORT ('OMMUNI('ATIONS
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lmV
D
I
I I
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2
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Fig. 2. Responses to two successive antidromic stimuli (upward arrows) at a critical interval. A, control; spike evoked by single stimulus. B and C, superimposed sweeps showing effect of first response (1) on response to second stimulus (2). D, E, F, single sweep records showing similar reactions in another neuron. Note change in latency, spike height and occasional block (F) of propagation beyond initial segment. See text.
Similar effects were produced on PVN cells by stimulation of the SON. Fig. 3G shows excitation of a PVN cell by SON stimulation and a subsequent inhibition of antidromic excitation. In H, SON stimulation which did not excite the PVN cell did not block its antidromic excitation. F is the control antidromic excitation. Similar occurrences are shown in a different pairing. Fig. 3I is an example of antidromic excitation of another PVN cell and J shows its orthodromic excitation from the SON. K illustrates what occurred when both stimuli were given in sequence and it can be seen that the effective SON stimulus inhibited the antidromic firing of the PVN cell. Inhibitory interaction was not confined to antidromic excitation. Rates of spontaneous firing in cells of one nucleus could be reduced by stimuli applied to the other. The mechanism of this interaction is under study. Not all but a significant number of SON and PVN cells (14 out of 35 cells tested) were found to be orthodromically excited or to have their antidromic excitation blocked from other neuroendocrine cells. Anatomical studies have shown that some nerve fibers from the PVN terminate in the SON 1. Our electrophysiological observations indicate that projections from the SON to the PVN also exist. The physiological implications of these interconnections require further study. Brain Research, 20 (1970)462-466
SHORT COMMUNICATIONS
465
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Fig. 3. Interaction between cells of SON and PV N. A D upward arrow indicates artefact of antidromic stimulation. I : A and D, spike evoked in SON cell by antidromic stimulation; B, C, and E, effects of PVN stimulation on response of SON neuron to antidromic stimulation. II: F, G, H and 1, J, K are from different neurons. F and I, response of PVN cell to antidromic stimulation; J, response of PVN cell to SON stimulation; G, H and K, effects of SON excitation on PVN response to antidromic stimulation. See text.
It has b e e n r e p o r t e d t h a t s e p a r a t e release o f a n t i d i u r e t i c h o r m o n e a n d o x y t o c i n d o e s o c c u r . T h i s i n h i b i t o r y i n t e r a c t i o n m a y f a c i l i t a t e the f u n c t i o n a l i n d e p e n d e n c e o f the two secretory processes.
This work was supported by U.S. Public Health Service Grant NB-6537-04 and NB-846-15. Department of Physiology, State University of New York, Downstate Medical Center, Brooklyn, N. Y. 11203 (U.S.A.)
HIROSHI YAMASHITA* KIYOMI KOIZUMI C H A N D L E R McC. BROOKS
1 BARGMANN,W., Ober die neurosekretorische Verkniipfung von Hypothalamus und Neurohypophyse, Z. Zellforsch., 34 (1949) 610-634. 2 BROOKS, C. McC., USmYAMA, J., AND LANGE, G., Reactions of neurons in or near the supraoptic nuclei, Amer. J. Physiol., 202 (1962) 487-490. 3 CROSS, B. A., NovlN, D., AND StmDSTAN, J. W., Antidromic activation of neurones in the paraventricular nucleus by stimulation in the neural lobe of the pituitary, J. Physiok (Lond.), 203 (1969) 68 P-69P. 4 DYBALL,R. E. J., AND KOZZUMI,K., Electrical activity in the supraoptic and paraventricular nuclei associated with neurohypophysial hormone release, J. Physiol. (Lond.), 201 (1969) 711-722. * Postdoctoral Fellow from the Department of Physiology, Kobe University, College of Medicine, Kobe (Japan).
Brain Research, 20 (! 970) 462-466
466
SHORT COMMUNICATIONS
5 ISHIKAWA,T., KOIZUMI, K., AND BROOKS,C. McC., Electrical activity recorded from the pituitary stalk of the cat, Amer. J. Physiol., 210 (1966) 427-431. 6 SUDA, I., KOIZUMI, K., ArxD BROOKS,C. McC., Study of unitary activity in the supraoptic nucleus of the hypothalamus, Jap. J. Physiol., 13 (1963) 374-385. 7 YAGI, K., AZUMA, T., AND MATSUOA,K., Neurosecretory cell: capable of conducting impulse in rats, Science, 154 (1966) 778.-779.
(Accepted April 6th, 1970)
Brain Research, 20 (1970)462-466
SHORT COMMUNICATIONS
Electrophysiological studies of neurosecretory cells in the cat hypothalamus The neuroendocrine cells of the hypothalamus are considered to perform a dual function in that they synthesize and secrete hormones as well as conduct nerve impulses. A number of authors have obtained electrophysiological evidence in support of this contention 3,4,7. Yagi e t al. 7 reported briefly that cells in the rat's supraoptic nucleus (SON) could be antidromicaily excited by stimulation of the posterior lobe of the pituitary. Dyball and Koizumi 4 found that neuroendocrine cells in the SON of the rat hypothalamus, which had been identified by antidromic stimulation of the posterior pituitary, were sensitive not only to osmatic changes but were also excited by stimulation of vagal afferent fibers and by injection of chemicals which increased antidiuretic hormone release. Cross e t al. 3 also studied antidromic excitation of the rat paraventricular nucleus (PVN) cells. The present report extends these studies to the cat hypothalamus. Despite previous failures to evoke antidromic excitation of the cells of the SON and PVN", 6 we finally succeeded in evoking responses from cells in these nuclei by single pulse (I msec duration) stimulation of the posterior lobe of the pituitary. The nature of these responses and interactions of the nuclei were studied in some detail. The following procedures were employed. Cats were anesthetized with chloralose (35-70 mg/kg) in most instances but occasionally with Nembutal (35 mg/kg). Fine bipolar silver or steel electrodes were imbedded in the posterior lobe of the pituitary after it had been exposed by hemispherectomy, or through the palate by removal of the sphenoid bone. Recordings were made from SON and PVN neurons by glass capillary electrodes filled with 3 M KCI or with a fast-green NaCI mixture. These electrodes were positioned stereotaxically, being introduced from the'ventral side of the hypothalamus, or visually placed from the hemispherectomized side. The precise positions of recording and stimulating electrode tips were determined histologically after each experiment. Some 34 experiments were performed and recordings obtained from 98 cells. Fig. I shows extracellular and intracellular recording of antidromic excitation of cells in the SON and PVN. These responses were evoked by single pulse stimulation of the posterior lobe of the pituitary. Latencies of the antidromic potentials varied as did the distance between stimulating and recording electrodes, but for each cell in a specific experiment latency was constant and did not fluctuate as in the case of orthodromic excitation. These latencies to antidromic stimulation fell between 10-20 msec for SON cells and 15-25 msec for PVN cells. By calculating average distances between stimulating and recording electrodes, it was estimated that conduction velocities in these neurosecretory cell axons ranged from 0.4 to 0.9 m/see. These values agree well with measurements made by others in rats and cats 3-'~.7. The antidromic potentials of the neurosecretory cells usually showed a notch in their rising phase indicative of delay in invasion of the soma by the axonal impulse. This notch became clearer when the stimulus intensity was low or when high frequency stimuli were given (Fig. 1A, C). Fig I B shows an intracellularly recorded antidromic potential. Brain Research,
20 (1970)462-466
463
SHORT COMMUNICATIONS
SON
PVN
1
mV
I mV $
i 5 msec
D
I0 msec Fig. 1. Antidromically induced action potentials recorded from the supraoptic nucleus (SON) (C and D) and paraventricular nucleus (PVN) (A and B). A and C show superimposed reactions obtained with barely threshold stimuli. Note the notch in the rising phase and that block occurred in some instances at this point. All records are from extracellular recordings except B which shows a spike induced during intracellular recording. Post-spike hyperpolarization was occasionally observed as in D.
When two successive stimuli were applied to the posterior lobe, the second response was blocked at 5-10 msec stimulus interval, the second evoking only a small response resembling an initial segment (IS) spike. This indicated that the refractory period of neurosecretory cells following antidromic excitation is of the order of 10 msec. Fig. 2 presents a series of records which show that at a critical stimulus interval the second of two antidromic responses has a longer latency (B, C, E), smaller amplitude (D, E) or is shown only as an IS spike (F). Full size uniform action potentials were evoked at stimulation rates of 50-100/sec. One unique result of this investigation was the discovery that cells of the SON and PVN interact in a rather complex fashion. It was found that some cells of the SON were excited orthodromically by pulses applied to PVN cells. Fig. 3C shows a response evoked immediately after PVN stimulation. Since recording was only from a SON cell it was the SON cell which gave the response. Interactions, however, were not invariably excitatory. It is shown in Fig. 3B, C and E that this same stimulation of the PVN inhibited antidromic excitation of the SON cell completely (B, C) or block propagation of the antidromic invasion at the initial segment (IS spike in E). Records A and D show control antidromic excitations of the SON cell under study. Brain Research, 20 (1970) 462-466
464
SHORT ('OMMUNI('ATIONS
~ 1 ~
A
] ImV
lmV
D
I
I I
l
I1
2
B
I
l
'
2
2
C
!
10m6ec 2
~t
2
Fig. 2. Responses to two successive antidromic stimuli (upward arrows) at a critical interval. A, control; spike evoked by single stimulus. B and C, superimposed sweeps showing effect of first response (1) on response to second stimulus (2). D, E, F, single sweep records showing similar reactions in another neuron. Note change in latency, spike height and occasional block (F) of propagation beyond initial segment. See text.
Similar effects were produced on PVN cells by stimulation of the SON. Fig. 3G shows excitation of a PVN cell by SON stimulation and a subsequent inhibition of antidromic excitation. In H, SON stimulation which did not excite the PVN cell did not block its antidromic excitation. F is the control antidromic excitation. Similar occurrences are shown in a different pairing. Fig. 3I is an example of antidromic excitation of another PVN cell and J shows its orthodromic excitation from the SON. K illustrates what occurred when both stimuli were given in sequence and it can be seen that the effective SON stimulus inhibited the antidromic firing of the PVN cell. Inhibitory interaction was not confined to antidromic excitation. Rates of spontaneous firing in cells of one nucleus could be reduced by stimuli applied to the other. The mechanism of this interaction is under study. Not all but a significant number of SON and PVN cells (14 out of 35 cells tested) were found to be orthodromically excited or to have their antidromic excitation blocked from other neuroendocrine cells. Anatomical studies have shown that some nerve fibers from the PVN terminate in the SON 1. Our electrophysiological observations indicate that projections from the SON to the PVN also exist. The physiological implications of these interconnections require further study. Brain Research, 20 (1970)462-466
SHORT COMMUNICATIONS
465
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Fig. 3. Interaction between cells of SON and PV N. A D upward arrow indicates artefact of antidromic stimulation. I : A and D, spike evoked in SON cell by antidromic stimulation; B, C, and E, effects of PVN stimulation on response of SON neuron to antidromic stimulation. II: F, G, H and 1, J, K are from different neurons. F and I, response of PVN cell to antidromic stimulation; J, response of PVN cell to SON stimulation; G, H and K, effects of SON excitation on PVN response to antidromic stimulation. See text.
It has b e e n r e p o r t e d t h a t s e p a r a t e release o f a n t i d i u r e t i c h o r m o n e a n d o x y t o c i n d o e s o c c u r . T h i s i n h i b i t o r y i n t e r a c t i o n m a y f a c i l i t a t e the f u n c t i o n a l i n d e p e n d e n c e o f the two secretory processes.
This work was supported by U.S. Public Health Service Grant NB-6537-04 and NB-846-15. Department of Physiology, State University of New York, Downstate Medical Center, Brooklyn, N. Y. 11203 (U.S.A.)
HIROSHI YAMASHITA* KIYOMI KOIZUMI C H A N D L E R McC. BROOKS
1 BARGMANN,W., Ober die neurosekretorische Verkniipfung von Hypothalamus und Neurohypophyse, Z. Zellforsch., 34 (1949) 610-634. 2 BROOKS, C. McC., USmYAMA, J., AND LANGE, G., Reactions of neurons in or near the supraoptic nuclei, Amer. J. Physiol., 202 (1962) 487-490. 3 CROSS, B. A., NovlN, D., AND StmDSTAN, J. W., Antidromic activation of neurones in the paraventricular nucleus by stimulation in the neural lobe of the pituitary, J. Physiok (Lond.), 203 (1969) 68 P-69P. 4 DYBALL,R. E. J., AND KOZZUMI,K., Electrical activity in the supraoptic and paraventricular nuclei associated with neurohypophysial hormone release, J. Physiol. (Lond.), 201 (1969) 711-722. * Postdoctoral Fellow from the Department of Physiology, Kobe University, College of Medicine, Kobe (Japan).
Brain Research, 20 (! 970) 462-466
466
SHORT COMMUNICATIONS
5 ISHIKAWA,T., KOIZUMI, K., AND BROOKS,C. McC., Electrical activity recorded from the pituitary stalk of the cat, Amer. J. Physiol., 210 (1966) 427-431. 6 SUDA, I., KOIZUMI, K., ArxD BROOKS,C. McC., Study of unitary activity in the supraoptic nucleus of the hypothalamus, Jap. J. Physiol., 13 (1963) 374-385. 7 YAGI, K., AZUMA, T., AND MATSUOA,K., Neurosecretory cell: capable of conducting impulse in rats, Science, 154 (1966) 778.-779.
(Accepted April 6th, 1970)
Brain Research, 20 (1970)462-466