Electrophysiology and pharmacology of projections from the suprachiasmatic nucleus to the ventromedial preoptic area in rat

doi:10.1016/S0306-4522(00)00170-6

Neuroscience

Volume 98, Issue 4, July 2000, Pages 715-728

https://doi.org/10.1016/S0306-4522(00)00170-6 Get rights and content

Abstract

Extracellular and whole-cell patch-clamp recordings were made from neurons in the ventromedial preoptic area in rat horizontal brain slices. Responses to single-pulse electrical stimulation of the ipsilateral suprachiasmatic nucleus were characterized using peristimulus time histograms or postsynaptic current recordings, and bath application of neurotransmitter receptor antagonists. Extracellular recordings showed that suprachiasmatic nucleus stimulation (50–150 μA) elicited a short-latency suppression in 35 of 64 neurons (55%), with the majority (29/35, 83%) showing a biphasic response consisting of a short-latency suppression followed by a long-duration activation. In addition, 14 cells (22%) showed activation only, while 15 (23%) were unresponsive. The GABA_A receptor antagonist bicuculline (5–10 μM) reversibly blocked suppressions evoked by suprachiasmatic nucleus stimulation (20/20 cells). In the majority of these neurons (13/20), bicuculline also unmasked an activation in response to stimulation. Activations elicited by suprachiasmatic nucleus stimulation, either in the presence or absence of bicuculline, were blocked by the non-N-methyl-d-aspartate and N-methyl-d-aspartate glutamate receptor antagonists 6,7-dinitroquinoxaline-2,3-dione and (±)-3-(2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (10/10 cells). 6,7-Dinitroquinoxaline-2,3-dione (10 μM) selectively and reversibly blocked the initial, short-duration (<50 ms) activation, but had no effect on the longer-duration activation. In contrast, (±)-3-(2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (10 μM) appeared to inhibit the long-duration activation selectively without affecting the initial rapid activation. Combined applications of the two ionotropic glutamate receptor antagonists blocked stimulation-induced activations completely. All the pharmacological effects were concentration dependent. Whole-cell patch-clamp recordings showed that suprachiasmatic nucleus stimulation elicited inhibitory postsynaptic currents or a combination of inhibitory and excitatory postsynaptic currents in 25 of 33 neurons tested. The inhibitory postsynaptic currents had short onset latencies (4.9±0.3 ms) and a reversal potential of −56.0±3.8 mV (n=5), and were reversibly blocked by bicuculline (5–10 μM, 4/4 cells). In the presence of bicuculline (5–10 μM), excitatory postsynaptic currents had short onset latencies (4.7±0.5 ms), and had a fast and a slow component. (±) 3-(2-Carboxypiperazin-4-yl)-propyl-1-phosphonic acid blocked the slow, but not the fast, component, whereas 6,7-dinitroquinoxaline-2,3-dione blocked the fast, but not the slow, component (n=7).

These results suggest that the projection from the suprachiasmatic nucleus conveys both inhibitory (GABA) and excitatory (glutamate) inputs to the ventromedial preoptic area. GABA_A receptor and both non-N-methyl-d-aspartate and N-methyl-d-aspartate glutamate receptors mediate these influences. These inputs may be responsible for conveying information related to circadian phase from the suprachiasmatic nucleus to regions of the preoptic area known to be involved in regulation of sleep/waking and other physiological functions.

Section snippets

Slice preparation

Data were obtained from male Wistar rats (Charles River Canada, St Constant, Québec) weighing 170–250 g for extracellular recordings and 70–150 g for whole-cell patch-clamp recordings. Animals were housed under a 12-h/12-h light–dark cycle with lights on at 07.00. All animals were handled in accordance with the guidelines of the Canadian Council on Animal Care. All efforts were made to minimize animal suffering and to reduce the number of animals used.

Rats were anesthetized with halothane and

Results

Data reported here were derived from a total of 24 hypothalamic slices for extracellular recording and 23 slices for whole-cell patch-clamp recording. Several VMPO neurons were recorded from each slice and characterized in terms of their responses to SCN stimulation. However, only one neuron in each slice was tested for the effects of antagonists.

Histological sections from two slices showing the general appearance of the slice and the locations of stimulation and recording sites are illustrated

Discussion

The extracellular recording results indicated that VMPO neurons respond to stimulation of the SCN with relatively short, fixed latencies, suggestive of a monosynaptic response. The whole-cell patch-clamp recording results showed that PSCs elicited by SCN stimulation had constant latencies that were independent of stimulation current and followed high-frequency stimulation (20 Hz) without fail. These results strongly suggest that SCN stimulation-evoked responses of VMPO neurons are mediated

Acknowledgments

This research was supported by the MRC (MT-14035) and Nova Scotia Department of Health. We thank Rob Mason for assistance in the design and construction of the recording apparatus, Steve Barnes, Charles Yang and Natalia Gorelova for technical advice and helpful comments on whole-cell patch-clamp recordings, Frank Smith for the use of a pipette puller, Joan Burns for assistance in performing histology, and Stephen Whitefield for assistance in digital imaging.

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Electrophysiology and pharmacology of projections from the suprachiasmatic nucleus to the ventromedial preoptic area in rat

Abstract

Section snippets

Slice preparation

Results

Discussion

Acknowledgments

Molec. Brain Res.

Neurosci. Lett.

Neurosci. Lett.

Brain Res.

Brain Res.

Neurosci. Lett.

Brain Res.

Regul. Pept.

Neuroscience

Neuron

Expl Neurol.

Brain Res.

Brain Res.

Neurosci. Lett.

Neurosci. Lett.

Expl Neurol.

Brain Res.

Prog. Brain Res.

Brain Res.

The neural pathway from the suprachiasmatic nucleus to the locus coeruleus: a transsynaptic tracing study

Soc. Neurosci. Abstr.

Ultrastructural evidence for intra- and extranuclear projections of GABAergic neurons of the suprachiasmatic nucleus

J. comp. Neurol.

Immunocytochemical localization of vasoactive intestinal polypeptide-containing cells and processes in the suprachiasmatic nucleus of the rat: light and electron microscopic analysis

J. Neurosci.

Glutamate-like immunoreactivity in retinal terminals of the mouse suprachiasmatic nucleus

Eur. J. Neurosci.

Sleep deprivation and c-fos expression in the rat brain

J. Sleep Res.

Neurones in the supraoptic nucleus of the rat are regulated by a projection from the suprachiasmatic nucleus

J. Physiol.

A simple insert for the PDMI-2 microincubator offers mechanical stability for brain slice recording

J. Physiol.

Human sleep: its duration and organization depend on its circadian phase

Science

GABA: a dominant neurotransmitter in the hypothalamus

J. comp. Neurol.