Elsevier

Neuropharmacology

Volume 45, Issue 5, October 2003, Pages 575-584
Neuropharmacology

Modulation of GABA release during morphine withdrawal in midbrain neurons in vitro

https://doi.org/10.1016/S0028-3908(03)00205-3Get rights and content

Abstract

Chronic treatment with opioids induces adaptations in neurons leading to tolerance and dependence. Studies have implicated the midbrain periaqueductal gray (PAG) in the expression of many signs of withdrawal. Patch-clamp recording techniques were used to examine whether augmentation of adenylyl cyclase signalling produces hyperexcitation in GABAergic nerve terminals within the mouse PAG. Both the rate of mIPSCs and the amplitude of evoked IPSCs during naloxone-precipitated withdrawal was profoundly enhanced in chronically morphine treated mice, compared to vehicle treated controls, in the presence but not the absence an adenosine A1 receptor antagonist DPCPX. Enhanced GABAergic transmission in the presence of DPCPX was abolished by blocking protein kinase A. Inhibitors of cAMP transport, phosphodiesterase and nucleotide transport mimicked the effect of DPCPX. Coupling efficacy of μ-receptors to presynaptic inhibition of GABA release was increased in dependent mice in the presence of DPCPX. The increased coupling efficacy was abolished by blocking protein kinase A, which unmasked an underlying μ-receptor tolerance. These findings indicate that enhanced adenylyl cyclase signalling following chronic morphine treatment produces (1) GABAergic terminal hyperexcitability during withdrawal that is retarded by a concomitant increase in endogenous adenosine, and (2) enhanced μ-receptor coupling to presynaptic inhibition that overcomes an underlying tolerance.

Introduction

Opioids have unrivalled clinical utility as analgesic agents although their use is limited by the development of tolerance and physical dependence (Koob and Le Moal, 1997). Physical dependence associated with chronic opioid use is characterised by a specific withdrawal syndrome that occurs after abrupt cessation of treatment or administration of an opioid receptor antagonist. Withdrawal from opioids results from counteradaptations developed in opioid-sensitive neurons and within the neuronal networks containing those neurons (Nestler and Aghajanian, 1997, Williams et al., 2001). Augmentation of the adenylyl cyclase → cyclic AMP (cAMP) → protein kinase A signalling cascade is a consistent cellular counteradaptation widely considered to be responsible for the expression of opioid withdrawal signs (Avidor-Reiss et al., 1995, Nestler and Aghajanian, 1997, Nestler and Tallman, 1988, Sharma et al., 1975).

The midbrain periaqueductal gray region (PAG) plays a key role in the expression of many of the somatic signs of opioid withdrawal (Bozarth, 1994, Bozarth and Wise, 1984, Chieng and Christie, 1996, Christie et al., 1997, Maldonado et al., 1992). Microinjection studies in the PAG have demonstrated that augmentation of the adenylyl cyclase → protein kinase A pathway many of these behavioural signs (Bozarth and Wise, 1984, Maldonado et al., 1995, Punch et al., 1997, Stinus et al., 1990, Wei et al., 1973).

A direct cellular consequence of hypertrophied adenylyl cyclase signalling is withdrawal induced protein kinase A-dependent enhancement of γ-aminobutyric acid (GABA)-mediated synaptic transmission in the PAG (Ingram et al., 1998), the ventral tegmental area (VTA) (Bonci and Williams, 1996, Bonci and Williams, 1997), the nucleus accumbens (NAcc) (Chieng and Williams, 1998) and the dorsal raphe (Jolas et al., 2000). Rebound activation of adenylyl cyclase during withdrawal is also proposed to account for the electrical hyperexcitability of opioid-sensitive PAG neurons (Chieng and Christie, 1996). Although hypertrophied adenylyl cyclase signalling is a general mechanism involved in augmented GABA-mediated synaptic transmission during opioid withdrawal (Shoji et al., 1999), the precise mechanisms regulating transmitter release may vary from region to region or from synapse to synapse. For example increased levels of endogenous adenosine resulting from enhanced cAMP formation plays a role in the rat NAcc (Chieng and Williams, 1998) and VTA (Bonci and Williams, 1996) but not in the rat PAG (Ingram et al., 1998). In the present study we provide novel information on the consequences of augmented adenylyl cyclase → cAMP → protein kinase A signalling during morphine withdrawal. Specifically we demonstrate that: (1) withdrawal-induced hyperexcitability in mouse PAG GABAergic terminals is mediated by augmentation of adenylyl cyclase signalling; (2) adenylyl cyclase-mediated hyperexcitation of PAG neurons is modulated by endogenous adenosine, acting on presynaptic A1 receptors and (3) upregulated cyclase activity mediates an increase in μ-receptor coupling to GABA release in dependent mice that overcomes an underlying tolerance present at the μ-receptor.

Section snippets

Methods

All animal experiments were conducted in accordance with National Institutes of Health guide for the care and use of laboratory animals.

Results

Male C57B16/J mice were treated chronically with either morphine to induce dependence, or vehicle alone to serve as controls (Chieng and Christie, 1996). This protocol induces intense naloxone-precipitated withdrawal behaviour as shown with similar studies in the rat (Ingram et al., 1998). Whole-cell voltage-clamp recordings of spontaneous GABAA mediated mIPSCs were made from PAG slices that were maintained in 5 μM morphine to prevent spontaneous opioid withdrawal. These spontaneous events were

Discussion

The present study demonstrated that during acute opiate withdrawal the probability of transmitter release from GABAergic terminals in the PAG of dependent mice is increased as a result of an upregulation of the adenylyl cyclase cascade. Augmentation of the adenylyl cyclase → cAMP → protein kinase A pathway during withdrawal had two major functional consequences. Firstly, there was an increase in efficacy of μ-receptor coupling to presynaptic inhibition of GABA release in opioid-dependent

Acknowledgements

This work was supported by the National Institute for Drug Abuse (DA12926-01) and The Medical Foundation of The University of Sydney.

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