Electrophysiology of the subfornical organ and its hypothalamic connections—an in-vivo study in the rat

Brain Research Bulierin, Vol. 15, pp. 85-86, 1985. * Ankho International Inc. Printed in

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Electrophysiology of the Subfornical Organ and its Hypothalamic Connectionsan In-Vivo Study in the Rat* L. P. RENAUD, A, V. FERGUSON,’ T. A. DAY,2 C. W. BOURQUE3 AND S. SGRO Neurosciences

Unit, The Montreal General Hospital Research Institute, and McGill University 1650 Cedar Avenue, Montreal, Quebec, Canada, H3G IA4

RENAUD, L. P., A. V. FERGUSON, T. A. DAY, C. W. BOURQUE AND S. SGRO. ~~ectrophysi~l~gy of the subf~rnjrai organ and its h~)pothalamjc c~nnectj~ns~n in-viva study in the rot. BRAIN RES BULL 15(l) 83-86, 1985.-The connections of the subfornical organ to a defined population of hypothalamic neurons have been explored with extracetlular recording methods in the rat. Electrical stimulation in the subfomical organ has a predominantly excitatory action on a majority of oxytocin and vasopressin-secreting neurosecretory neurons in the supraoptic and paraventricular nuclei. Subfornical organ stimulation also enhances the excitability of paraventricular nucleus neurons that project to the median eminence, and to the dorsomedial medulla. These observations provide initial evidence of functional connectivity of subfomical organ neurons with other hypothalamic cells that are engaged in central regulation of pituitary secretions and autonomic activities. Subfomicai organ

Hypothalamus

Neurosecretory

neurons

THE efferent connections of the subfomical organ (SFO) have now been defined by various anatomical tracer methods [9, 10, 13, 16, 191 and leave little doubt that the supraoptic (SON) and paraventricular (PVN) nuclei of the hypothalamus are prominent sites where SF0 neurons are likely to exert their influence. These nuclei contain neurons with major homeostatic functions, e.g., regulation of anterb and posterior pituitary hormone release and various autonomic activities [19]. Studies reviewed by other participants in this workshop attest to the SFO’s involvement in body fluid balance and cardiovascular regulation. In this article we review electrophysiological studies on the SF0 and the interrelationships between the SF0 and a select population of hypothalamic neurons involved in regulatory functions.

Oxytocin

Vasopressin

In vivo studies show that iontophoretically applied AI1 results in the excitation of single SF0 neurons 12,141. Small reductions in blood pressure have aIso been shown to cause repeatable increases in the firing frequency of single SF0 neurons [8,14]. OXYTOCIN

(OXY) AND VASOPRESSIN

(VP) SECRETING

NEURONS

Magnocellular neurosecretory neurons in the SON and PVN are known to synthesize either OXY or VP 120) and to project to the posterior pituitary [ 181. Electrophysiological single unit recording techniques permit the identification of this group of cells according to their antidromic responses to stimulation in the posterior pituitary; further clues as to the chemical identity of these cells derives from both their spontaneous activity patterns and their responses to baroreceptor activation [7,15]. Thus, phasically and continuously active cells inhibited by baroreceptor activation may be classified as putative VP-secreting neurons while continuously firing cells unaffected by baroreceptor activation are classified as putative OXY-secreting cells. We have examined the effects in vivo of electrical stimulation in the SF0 on the excitability of such “identified” oxytocin and vasopressin containing neurons [S, 171. The predominant effect observed in response to a single stimulus delivered to the SF0 was a relatively long latency (ca. 80 msec) and prolonged (up to 500 msec) increase in excitability (Fig. 1). Shorter latency and briefer initial excitations, or initial reductions in excitability (alone, or followed by a later excitation) have also been observed but with less frequency (Fig. 2). These effects were intensity

SF0 NEURONS

The effects of specific physiological and pharmacological perturbations on the activity of SF0 neurons have been examined in a limited number of in vivo and in vitro studies. Buranarugsa and Hubbard [l] have recorded extracellular action potentials from SF0 slices and shown single units to be activated by both ~giotensin II (AII) and carbachol. They suggest that the AI1 sensitive units may serve as interneurons within the SF0 projecting to the carbachol sensitive neurons which then form the major efferent projections from the SFO. However, double labelling studies [9] have now demonstrated that at least a part of the efferent projections of the SF0 consist of AI1 containing neurons.

*From the Workshop The S~bfornica~ Organ as a Model of Neur~humorai for Neurosciences, Anaheim, CA, October 15, 1984.

Integration,

presented at the 14th Annual Meeting of the Society

‘Current address: Queen’s University, Dept. of Physiology, Kingston, Ontario. ‘Current address: University of Western Ontario, Dept. of Physiology, London, Ontario. 3Current address: School of Pharmacy, Brunswick Square, London, England.

83

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RENAUD

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PVN (n-107)

200 !A

0.9

T 43

13 2

14

2f

2

21

13

L

13

3

5

18

13

21

E

4OOpA

1.2T

L

FIG. 2. Summary schema of SF0 effects on neurohypophysealprojecting cells classified according to their location (SON or PVN). chemical identity (VP or OXY-secreting) and their patterns of response observed in poststimulus histograms (centre column; arrow depicts SF0 stimulus; horizontal bar is 100 msec). The latter include (A) long duration excitability increase, (B) depression followed by a long duration excitation, (C) depression only, (D) short duration increase in excitability and (E) no response.

inte~elationships in the control of body fluid balance awaits further investigation. 600 mroc

FIG. 1. Poststimulus

histograms obtained from a putative VPsecreting SON neuron illustrate (on the left) an intensity dependent long duration increase in excitability following a single SF0 stimulus at the parameters indicated, and (on the right) the response to a subthreshold (0.9xT) and a snprathreshold (1.2xT) stimulus in the posterior pituitary. Each histogram contains 200 sweeps. Note that the increase in excitability evoked by the SF0 stimulus is preceded by a brief reduction in firing.

dependent (Fig. 1) and site specific to the SFO, and involved both putative OXY- and VP-secreting neurons in both the SON and PVN (Fig. 2). Moreover, the differing types of response could all be elicited from a single SF0 stimulation site indicating some degree of heterogeneity in the SF0 projections to these neurosecretory cells (cf. Fig. 2 in 1171). While a direct (monosynaptic) pathway from SF0 to SON and PVN is implied on the basis of light [10,13] and electronmicroscopic [ 161 data, recent electraphysiological observations have indicated the probability of an oligosynaptic pathway from SF0 to these neurosecretory cells. Thus, approximately 60% of neurons in the medial septum/diagonaI band of Broca (MS-DBB) region that can be ~tidrorn~~~Iy identified as projecting to the region of the SON display orthodromic excitation following an electrical stimulus in SFO; a reduction in excitability was seldom observed [3]. A smaller number of cells in this region (8%) could be antidromically identified as projecting to the SFO, while 3 neurons were antidromically activated from both SF0 and SON, indicating their simul~neous projection to both of these regions. The role of these SFO-MS-DBB and SON

TUBEROINFUNDIBULAR NEURONS

Although effects of SF0 stimulation were observed on neurohypophyseal-projecting OXY- and VP-secreting cells in both the SON and PVN, a large proportion (approximately SG%) of PVN neurons which did not project to the posterior pituitary were also found to be o~hodromically activated by SF0 stimulation. A number of such neurons have been classified as tuberoinfundibular cells on the basis of their antidromic activation from the median eminence (Fig. 3). Of 55 tu~roinfundibul~ cells tested, 20% showed short latency short duration increases in excitability in response to single SF0 stimuli f5]. Although the chemical identity of this group of tuberoinfundibular cells remains undefined, they are most likely to contain either TRH or CRF, the two most numerous PVN cell groups which project to the median eminence. CAUDALLY PROJECTING PVN NEURONS

There is now considerable evidence demonstrating that either electrical or chemical stimulation in the SF0 elevates arterial blood pressure [8, 11, 121. We have recently reported [6] that such increases in blood pressure in response to electrical stimulation in the SF0 are greatly reduced by bilateral electrolytic lesions of the PVN. thus implicating SF0 efferents to this region in the pressor response. The short latency and rapid off response indicate the effect to be neurally rather than humorally mediated, possibly through descending PVN projections to the medulla and/or spinal cord. We have also observed 141 that SF0 stimulation enhances the excitability of PVN neurons antidromically identified as projecting to the dorsomedial medulla, in the area of both the nucleus tractus solitarius and the dorsal motor nucleus of the vagus. Of this group of neurons, some !X% of spontaneously active units were activated by SF0 stimulat;on (Fig. 41. Both

ELECTROP~YSIOLOGY

85

OF THE SF0

neuron. On the left, oscilloscope sweeps illustrate all-or-none constant latency response to a single stimulus delivered to the dorsomedial medulla (DM; upper trace), constant latency responses to two suprathreshold stimuli (middle trace) and collision cancellation of the first antidromic spike by a spontaneous action potential (*) that occurs within the critical interval. On the right, poststimulus histograms (X axis maximum 1000 msec, Y axis maximum 20 counts per bin, resolution 4 msec per bin, 200 sweeps each) depict a variable latency increase in excitability evoked by SF0 stimuIation at two different stimulus strengths, and a constant latency (antidromic) response to DM stimulation. (From 141, with permission).

EMtNENCE

FIG. 3. Oscilloscope records obtained from a PVN tuberoinfundibuJar ceil. In the top record, superimposed sweeps demonstrate constant latency responses following two stimuli presented on the surface of the median eminence (ME; arrows refer to shock artifacts). In the middle trace, a single orthodromically generated action potential follows stimulation in the SFO; this spike lies outside the critical period for cancellation of the antidromic spike from a ME stimulus. In the lower record, three superimposed traces illustrate that each SFO-evoked spike (note the variable latencies indicating their orthodromic nature) is timed to fall within the critical period for cancellation of the ME-evoked antidromic response (from [5], with permission).

short and longer latency effects were observed, occasionalfy on a single neuron. Whether this projection represents the effector pathway mediating SFO-induced cardiovascular responses is not as yet clear; PVN projections to other spinal autonomic regions, e.g., the intermediolateral cell column, may also be involved. CONCLUSIONSANDCOMMENT

In-vivo electrophysiological studies of certain efferent projections of the SF0 have now provided the initial information on functional connectivity that can be useful in the

DRINKING BEHAVIOR VASOPRESSIN

AUTONOMIC

AOENOHYPOPHYSIS

CENTERS IBLOOD

PRESSURE)

FIG. 5. Schematic diagram depicts some SF0 connections with the hypothalamus that can be examined by electrophysiology. Also indicated are functional roles of these hypothalamic neurons proposed on the basis of their axonal projections to the neurohypophysis (VP and OXY-secreting), median eminence (regulation of anterior pituitary hormone release) and dorsomedial medulla (? cardiovascular regulation). Abbreviations: CSF, cerebrospinal fluid; MPG, medial preoptic area: MS, medial sentum: DBB, diagonal band of Broca; PVl6, hypothalamic paraventhcular nuclei; SON, hy~thalamic supraoptic nuclei; AV3V, anteroventral aspect of the third ventricle; VP, vasopressin; OXY, oxytocin.

evaluation of the role of this stucture in controlling the excitability of different hypothalamic cell groups involved in the control of a variety of homeostatic functions (Fig. 5). While SF0 neurons respond to chemical changes within their local environment, it would appear that activation of SF0 efferents has a predominantly excitatory influence on (I) the OXY and VP neurons of the SON and PVN; (2) PVN neurons projecting to the dorsomedial medulla; (3) PVN

86

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neurons projecting to the median eminence, and (4) neurons in the MS-DBB projecting to the region of the SON. The neurotransmitters involved in these pathways have not heen determined. However, the demonstration that some of these are AI1 projections suggests the involvement of at least this peptide and provides an intriguing area for future neuropharmacological study.

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ACKNOWLEDGEMENTS

Supported by MRC of Canada, AHFMR and FRSQ. Thanks to Gwen Peard for typing the manuscript, and to Fran&e Renaud and Joanne Rogers for technical assistance.

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