Elsevier

Neuropharmacology

Volume 52, Issue 2, February 2007, Pages 527-535
Neuropharmacology

Pre-synaptic histamine H3 receptors regulate glutamate, but not GABA release in rat thalamus

https://doi.org/10.1016/j.neuropharm.2006.08.001Get rights and content

Abstract

We have investigated the presence of histamine H3 receptors (H3Rs) on rat thalamic isolated nerve terminals (synaptosomes) and the effect of their activation on glutamate and GABA release. N-α-[methyl-3H]histamine ([3H]-NMHA) bound specifically to synaptosomal membranes with dissociation constant (Kd) 0.78 ± 0.20 nM and maximum binding (Bmax) 141 ± 12 fmol/mg protein. Inhibition of [3H]-NMHA binding by histamine and the H3R agonist immepip fit better to a two-site model, whereas for the H3R antagonist clobenpropit the best fit was to the one-site model. GTPγS (30 μM) decreased [3H]-NMHA binding by 55 ± 4% and made the histamine inhibition fit better to the one-site model. Immepip (30 nM) induced a modest, but significant increase (113 ± 2% of basal) in [35S]-GTPγS binding to synaptosomal membranes, an effect prevented by clobenpropit (1 μM) and by pre-treatment with pertussis toxin. In thalamus synaptosomes depolarisation-induced, Ca2+-dependent glutamate release was inhibited by histamine (1 μM, 25 ± 4% inhibition) and immepip (30 nM, 38 ± 5% reduction). These effects were reversed by clobenpropit (1 μM). Conversely, immepip (up to 1 μM) had no effect on depolarisation-evoked [3H]-GABA release. Extracellular synaptic responses were recorded in the thalamus ventrobasal complex by stimulating corticothalamic afferents. H3R activation reduced by 38 ± 7% the glutamate receptor-mediated field potentials (FPs), but increased the FP2/FP1 ratio (from 0.86 ± 0.03 to 1.38 ± 0.05) in a paired-pulse paradigm. Taken together, our results confirm the presence of H3Rs on thalamic nerve terminals and show that their activation modulates pre-synaptically glutamatergic, but not GABAergic neurotransmission.

Introduction

The thalamus links the brainstem, basal ganglia and cerebral cortex, and also serves as a relay transferring sensory information from the periphery and motor information originated in the cerebellum to the neocortex (Sommer, 2003). Thalamic function is mostly based on excitatory glutamatergic transmission and inhibitory GABAergic modulation, with serotonin, noradrenaline and histamine also playing significant roles in modulating thalamocortical functions (McCormick and Bal, 1997).

The cell bodies of histaminergic neurones are located exclusively in the tuberomammillary nucleus of the hypothalamus, from where they send diffuse projections to practically all brain regions (Wada et al., 1991). Histamine regulates pre- and post-synaptically a number of brain functions, through interaction with G protein-coupled receptors. Four such receptors (H1–H4) have been cloned, and three of them (H1, H2, and H3) are widely distributed in the mammalian central nervous system (Brown et al., 2001, Haas and Panula, 2003). The thalamus is innervated by histaminergic fibres (Panula et al., 1989, Jin et al., 2002), expresses H1, H2 and H3 receptors as well as their corresponding mRNAs (Vizuete et al., 1997, Jin et al., 2002, Jin et al., 2005), and there is evidence that neuronal histamine regulates thalamic functions through the activation of H1 and H2 receptors. For instance, in cat and guinea-pig thalamus the firing pattern of relay neurones is modulated by H1 receptors via the reduction of an ionic current identified as a “leak” K+ conductance (IKleak or IKL), and by H2 receptors by enhancing the hyperpolarisation-activated cationic current (Ih) or activating a chloride conductance (McCormick and Williamson, 1991, McCormick, 1992, McCormick and Bal, 1997, Lee et al., 2004).

In human thalamus mRNA encoding for H3 receptors (H3Rs) displayed the highest expression intensity among all three histamine receptors (Jin et al., 2002); however, little is known regarding their functional effects. The H3R was discovered originally on histamine containing neurones as a pre-synaptic receptor regulating the release and synthesis of histamine, but also regulates the release of other neurotransmitters/modulators such as acetylcholine, dopamine, noradrenaline, serotonin, GABA and glutamate (reviewed by Hill et al., 1997, Brown et al., 2001, Haas and Panula, 2003, Leurs et al., 2005).

H3Rs show a highly heterogeneous distribution in rat thalamus (Pillot et al., 2002, Jin et al., 2005) and the discrepancies between receptor levels and the expression of the corresponding mRNA suggest that at least in some nuclei H3Rs correspond to pre-synaptic receptors. These receptors may be located on the axon terminals of neurones located in cortical layers V and/or VI that use glutamate as neurotransmitter and express high levels of H3R mRNA (Pillot et al., 2002). In addition, the high expression of mRNA and very low expression of H3Rs in the reticular nucleus suggests that H3Rs are also located on the nerve terminals of reticular GABAergic neurones.

We report herein a study of the pharmacological characteristics of H3Rs present on isolated nerve terminals (synaptosomes) from rat thalamus, and the effect of their activation on glutamate release (detected by a fluorometric assay and by electrophysiological recording) and on [3H]GABA release from superfused synaptosomes.

Section snippets

Animals

Rats (males, Wistar strain), bred in the Cinvestav facilities, were used through out. Animals were maintained under constant room temperature (23 °C) and light/dark cycle (12:12 h), with food and water ad libitum. All procedures were in accordance to the “Guide for the care and use of laboratory animals” of the Mexican Council for Animal Care and approved by the Cinvestav Animal Care Committee. All efforts were made to minimise animal suffering and to use only as many animals were required for

[3H]-NMHA binding

Specific [3H]-NMHA binding to membranes from rat thalamus synaptosomes (Fig. 1A) yielded maximum binding (Bmax) 141 ± 12 fmol/mg protein (four experiments). The estimate for the equilibrium dissociation constant (Kd) was 0.78 ± 0.20 nM, in good agreement with values reported for rat cerebral cortex (0.7 nM, Kathmann et al., 1993), mouse cerebral cortex (0.71 nM, Nickel et al., 2001) and guinea-pig brain (0.4 nM, Korte et al., 1990). [3H]-NMHA binding was inhibited in a concentration-dependent manner by

Discussion

H3Rs are expressed in the thalamus with a highly heterogeneous distribution that in the rat roughly parallels histaminergic innervation, but does not match the distribution of the corresponding mRNA (Jin et al., 2002, Pillot et al., 2002). This discrepancy suggests that at least in some thalamic nuclei there exist pre-synaptic H3Rs, most likely located on corticothalamic glutamatergic terminals and GABAergic axons projecting from the reticular nucleus. We show herein that rat thalamic nerve

Conclusions

The thalamus is critical for information processing and integrative functions, and corticothalamic information appears to bear particular relevance to select input signals according to the requirements of cortical processing, and to enhance the synchronisation of oscillating cells during states of vigilance (McCormick and Bal, 1997, Jones, 2002). Our results indicate that, in addition to post-synaptic H1 and H2 receptors, H3Rs contribute to regulating thalamic function through the pre-synaptic

Acknowledgements

Supported by Cinvestav, Conacyt (grant 37345-N) and Third World Academy of Sciences. This work is based on the thesis submitted by B. Garduño-Torres to obtain the M.Sc. degree (Neurobiology) from Cinvestav. The excellent technical assistance of J.J. Sierra is gratefully acknowledged. B.G.-T. was the recipient of a Conacyt predoctoral scholarship.

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