Elsevier

European Journal of Pharmacology

Volume 767, 15 November 2015, Pages 144-151
European Journal of Pharmacology

Neuropharmacology and analgesia
Excitatory drive from the Subthalamic nucleus attenuates GABAergic transmission in the Substantia Nigra pars compacta via endocannabinoids

https://doi.org/10.1016/j.ejphar.2015.09.050Get rights and content

Abstract

Endocannabinoids (eCBs) are cannabis-like substances produced in the brain where their primary function is to regulate synaptic transmission by inhibiting neurotransmitter release in a retrograde fashion. We have recently demonstrated a novel mechanism regulating GABAergic transmission from neurons in the Substantia Nigra pars reticulata (SNr) to dopaminergic neurons in the Substantia Nigra pars compacta (SNc) mediated by eCBs. Production of eCBs was initiated by spillover of glutamate, yet the source of the glutamate was not determined (Freestone et al., 2014; Neuropharmacology 79 p467). The present study aimed at elucidating the potential role of glutamatergic terminals arising from neurons in the Subthalamic nucleus (STN) in driving the eCB-mediated modulation of this inhibitory transmission. GABAergic IPSCs or IPSPs evoked in SNc neurons by electrical stimuli delivered to the SNr region were transiently inhibited by electrical or pharmacological (U-tube application of muscarinic agonist carbachol [100 µM]) stimulation of the STN (to 74±5% and 69±4% respectively). In both stimulation protocols, the attenuation of GABAergic transmission was abolished by cannabinoid receptor 1 antagonist rimonabant (3 µM), and reduced by group 1 metabotropic glutamate receptor antagonist CPCCOEt (100 µM), consistent with a glutamate-initiated and eCB-mediated mechanism. The carbachol-induced attenuation of GABAergic transmission was abolished by M3 muscarinic receptor antagonist 4-DAMP (10 µM), confirming a specific activation of STN neurons. These results demonstrate that glutamatergic projection from the STN to dopaminergic SNc neurons underlies an eCB-mediated inhibition of GABAergic input to these neurons.

Introduction

The endocannabinoid (eCB) system comprises of CB1 cannabinoid receptors predominantly located on presynaptic terminals and endogenous ligands (endocannabinoids, eCBs) produced in postsynaptic neurons which, by acting on CB1 receptors, attenuate neurotransmitter release (Kano et al., 2009). Thus, the primary role of eCBs is modulation of synaptic transmission in a retrograde and local fashion. While the eCB system has been the topic of extensive studies since its key components have been discovered in 2001 (Kreitzer and Regehr, 2001, Ohno-Shosaku et al., 2001, Wilson and Nicoll, 2001), this work has been conducted mainly in the hippocampus and cerebellum, with relatively few studies conducted in the basal ganglia, particularly in ventral midbrain which shows high expression of CB1 receptors (Herkenham et al., 1991, Herkenham et al., 1990). The basal ganglia is a network involved in control of a variety of functions including voluntary movement and learning, modulated by dopamine-producing neurons of the Substantia Nigra pars compacta (SNc). Despite the rich expression of CB1 the role of eCBs in this brain region has received little attention (Wallmichrath and Szabo, 2002).

We have recently shown that GABAergic transmission from the Substantia Nigra pars reticulata (SNr; a major output nucleus of the basal ganglia) onto SNc dopaminergic neurons is attenuated by eCBs, and that this modulation is initiated by glutamate (Freestone et al., 2014). However, the source of this glutamatergic input has not been identified. The Subthalamic nucleus (STN) has significant glutamatergic projections to the ventral midbrain, including nigral dopaminergic neurons (Watabe-Uchida et al., 2012), and we now hypothesize that this nucleus is a source of glutamate driving the eCB modulation that we have observed. The STN is part of the classic ‘indirect’ pathway of the basal ganglia, relaying motor commands from the motor cortex-striatum-external Globus Pallidus circuit (Gerfen and Surmeier, 2011). In addition, STN forms a part of the ‘hyperdirect pathway’ relaying information directly from the cortex to SNr neurons (Bosch et al., 2012, Nambu et al., 2002). The present study, conducted in acute rat brain slices, investigated the role of the STN neurons in the eCB-mediated modulation of GABAergic transmission from the SNr to SNc.

Section snippets

Animal handling and acute slice preparation

All procedures were approved by the animals ethics committee of the University of Auckland and in accordance with New Zealand Government Animal Welfare Act. Brain slices were prepared from Wistar rats (P15-19) as described elsewhere (Freestone et al., 2009, Freestone et al., 2014). Briefly, brains were quickly removed from anaesthetized rats and placed in carbogen (95% O2/5% CO2)-bubbled chilled artificial cerebrospinal fluid (ACSF) containing (mM): NaCl 126, NaHCO3 24, glucose 10, KCl 2.5, CaCl

Results

Whole-cell current and voltage clamp recordings were made from 76 dopaminergic SNc neurons (Rm 126±5 MΩ, Cm 105±5 pF, Vm −56.4.±0.8 mV) which had relatively large and multipolar cell bodies (20–30 µm) and fired action potentials at low frequency (1.4±0.2 Hz). To confirm correct placement of the stimulating electrode in the STN, additional whole-cell recordings were made from STN neurons which showed properties consistent with those described in previous studies (Gajendiran et al., 2005, Nakanishi et

Discussion

We investigated the role of the STN in modulating synaptic transmission in downstream dopaminergic neurons in the SNc. Our results show that electrical stimulation of glutamatergic STN neurons reduced GABAergic transmission from SNr neurons to dopaminergic SNc neurons and that this effect is mediated by eCBs. The degree of GABAergic attenuation was proportional to the extent of STN activation. In addition, STN neurons were also stimulated using local U-tube application of muscarinic

Acknowledgments

This study was supported by the Auckland Medical Research Foundation (to PF) and Aotearoa Foundation Centre for Brain Research Fellowship (to PF).

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