Adaptive changes in the number of Gs- and Gi-proteins underlie adenylyl cyclase sensitization in morphine-treated rat striatal neurons

doi:10.1016/0922-4106(93)90165-6

European Journal of Pharmacology: Molecular Pharmacology

Volume 245, Issue 1, 15 March 1993, Pages 23-29

https://doi.org/10.1016/0922-4106(93)90165-6 Get rights and content

Abstract

In the present study we investigated the possible role of changes in the number of membrane-bound G-proteins in the sensitization of dopamine D₁ receptor-stimulated adenylyl cyclase, observed in primary cultures of rat striatal neurons chronically exposed to morphine. Whereas exposure of these neurons to 10 μM morphine for 7 days caused a profound increase in cyclic AMP production, induced by the dopamine D₁ receptor agonist SKF 38393 (1 μM), Scatchard analysis of [¹²⁵I]SCH 23982 binding to membrane preparations revealed that neither the B_max nor the K_d values of dopamine D₁ receptor binding sites were affected. Interestingly, immunoblotting experiments revealed an increase (of more than 50%) in the number of stimulatory G-proteins (G_αs) in neurons displaying an enhanced adenylyl cyclase activity. In morphine-treated neurons, the number of inhibitory G-proteins (G_αi) appeared to be slightly reduced (by about 16%). Moreover, the observation that cholera toxin (0.1 nM)-stimulated cyclic AMP production, unlike that induced by forskolin (1 μM), was enhanced in morphine-treated neurons, indicates a causal relationship between the reciprocal changes in G-protein number and the increase of dopamine D₁ receptor-stimulated adenylyl cyclase activity. The possible role of these changes in G-protein number in the development of morphine tolerance and dependence is discussed.

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Temporal and anatomic patterns of immediate-early gene expression in the forebrain of C57BL/6 and DBA/2 mice after morphine administration
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Citation Excerpt :
The induction may be a consequence of the previous direct inhibitory action of morphine on striatal neurons (cf Section “Saline injection effects on IEG expression and the inhibition of these effects by morphine”), to which cellular adaptations develop, and these adaptations are uncovered when the action of morphine ends. Sensitization of the adenylate cyclase – cyclic adenosine monophosphate (cAMP) – protein kinase A (PKA) signaling is the best-known cellular adjustment to long-term opioid exposure, and in striatal neurons, this sensitization can be observed as early as after 2 h of constant exposure to morphine (Duman et al., 1988; Van Vliet et al., 1991, 1993; Cao et al., 2010). Because the brain concentration of morphine peaks at 0.5 h after its peripheral administration to the mouse and because the half-life of the drug is approximately 1 h (Ishikawa et al., 1982, 1983; Barjavel et al., 1995), biologically relevant concentrations of the opioid in the brain may persist for a few hours.
Although morphine was previously reported to produce an instant induction of c-fos in the striatum, our recent studies have demonstrated that the expression of numerous immediate early genes (IEGs) is significantly elevated at delayed time-points (several hours) after morphine administration. To better dissect the time-course of opioid-produced IEG induction, we used in situ hybridization to examine the expression of the IEGs c-fos, zif268 and arc in the mouse forebrain at several time-points after acute morphine injection. To link drug-produced behavioral changes with the activity of specific neuronal complexes, this study was performed comparatively in the C57BL/6 and DBA/2 mouse strains, which differ markedly in their locomotor responses to opioids and opioid reward.
Our study demonstrates that morphine produces two episodes of IEG induction, which are separate in time (30 min vs. 4–6 h) and which have different neuroanatomic distribution. At 30 min, one or more IEGs were induced in circumscribed subregions of the dorsal striatum (dStr) and of the nucleus accumbens (NAc) shell, as well as in the lateral septum. The observed inter-strain differences in IEG expression at 30 min support earlier proposals that activation of the dorsomedial striatum may mediate morphine-elicited locomotor stimulation (both effects were present only in the C57BL/6 strain). In contrast, NAc shell activation does not appear to be linked to morphine-elicited changes in locomotor behavior. The second IEG induction (of arc and of zif268) was more widespread, involving most of the dStr and the cortex. The second IEG induction peaked earlier in the DBA/2 mice than in the C57BL/6 mice (4 h compared with 6 h) and displayed no apparent relation to locomotor behavior. This delayed episode of IEG activation, which has largely been overlooked thus far, may contribute to the development of long-term effects of opioids such as tolerance, dependence and/or addiction.
Neural mechanisms of reproduction in females as a predisposing factor for drug addiction
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What is clear is that this increase in synaptic dopamine concentration enhances post-synaptic dopamine signaling. It has long been known that drugs of abuse not only sensitize the mesolimbic dopamine system but also alter receptor-mediated intracellular signaling [179,188,189,208,211,213]. In an effort to resolve components of intracellular sensitization that result from sexual experience, we measured cAMP accumulation in homogenates from the nucleus accumbens of female hamsters after in vitro pharmacological stimulation [22].
There is an increasing awareness that adolescent females differ from males in their response to drugs of abuse and consequently in their vulnerability to addiction. One possible component of this vulnerability to drug addiction is the neurobiological impact that reproductive physiology and behaviors have on the mesolimbic dopamine system, a key neural pathway mediating drug addiction. In this review, we examine animal models that address the impact of ovarian cyclicity, sexual affiliation, sexual behavior, and maternal care on the long-term plasticity of the mesolimbic dopamine system. The thesis is that this plasticity in synaptic neurotransmission stemming from an individual’s normal life history contributes to the pathological impact of drugs of abuse on the neurobiology of this system. Hormones released during reproductive cycles have only transient effects on these dopamine systems, whereas reproductive behaviors produce a persistent sensitization of dopamine release and post-synaptic neuronal responsiveness. Puberty itself may not represent a neurobiological risk factor for drug abuse, but attendant behavioral experiences may have a negative impact on females engaging in drug use.
Dopamine D<inf>1</inf> receptor-dependent modifications in the dopamine and cAMP-regulated phosphoprotein of M<inf>r</inf> 32 kDa phosphorylation pattern in striatal areas of morphine-sensitized rats
2009, Neuroscience
Morphine sensitization is a model of latent, functionally inducible increase in dopamine D₁ receptor–mediated transmission, which may be unmasked by an external stimulus. Morphine-sensitized rats present dopamine D₁ receptor–dependent stereotypies upon morphine challenge and resilience to unavoidable stress-induced behavioral deficits. This tonic increase in dopamine D₁ dopaminergic transmission is counter-adaptive to an enhanced μ-opioid receptor-dependent signaling in striatal areas. Control and sensitized rats show a similar dopamine and cAMP-regulated phosphoprotein of M_r 32 kDa (DARPP-32) phosphorylation pattern in striatal areas. Acute morphine administration induced an early increase and delayed decrease in phospho-threonine (Thr)34 DARPP-32 levels accompanied by a delayed increase in phospho-Thr75 DARPP-32 levels in the nucleus accumbens and caudate-putamen of sensitized rats, while it had no effects in control animals. The administration of a selective dopamine D₁ receptor antagonist (SCH 23390) before morphine challenge prevented the behavioral and neurochemical modifications in sensitized rats. 6-Methyl-2-(phenylethynyl)-pyridine, a selective metabotropic glutamate receptor 5 (mGluR₅) antagonist, administered 1 h after morphine challenge, prevented the delayed phosphorylation changes, but it had no effect when administered before challenge. Moreover, the DARPP-32 phosphorylation pattern in the caudate-putamen of sensitized rats after unavoidable stress exposure was studied. The stress-induced neurochemical modifications and their sensitivity to receptor antagonists were similar to those observed after acute morphine administration. In conclusion, these results suggest that in the experimental conditions used an increase in dopamine output in striatal areas is followed by a complex neurochemical pattern, in which the initial stimulation of dopamine D₁ receptors triggers a sequence of signaling events that lead to an mGluR₅-mediated increase in phospho-Thr75 DARPP-32 levels. Since DARPP-32 phosphorylated in Thr75 inhibits cAMP-dependent protein kinase (PKA) activity, the final result is a decrease in the dopamine D₁ receptor–dependent phosphorylation events.
Dexamethasone mimics the inhibitory effect of chronic pain on the development of tolerance to morphine analgesia and compensates for morphine induced changes in G proteins gene expression
2006, Brain Research
Citation Excerpt :
Opioid receptors are coupled to their effectors through inhibitory guanosine nucleotide binding proteins (G proteins) (Williams et al., 1988). Uncoupling of opioid receptors and G proteins (Liu and Anand, 2001), changes in the G protein content of striatal neurons (Vliet et al., 1993) and increased immunoreactivity of the α subunits of Gi/o proteins in Locus ceruleus (Nestler, 1997) are reported following chronic morphine administration. We previously reported that chronic administration of morphine increased the expression of β subunits of G proteins in lumbar spinal cord (Javan et al., 2005).
It is previously reported that the HPA axis plays role in the inhibitory effect of pain on tolerance development to analgesic effect of opioids. The present study was designed to investigate whether the chronic co-administration of dexamethasone as a glucocorticoid is also able to prevent or reverse analgesic tolerance to morphine and to compare the expression of G_αi/o and G_β subunits of G proteins in the context of chronic dexamethasone, development of morphine tolerance and their combination.
Analgesic tolerance to morphine was induced by chronic intraperitoneally (i.p.) administration of morphine 20 mg/kg to male Wistar rats weighing 200–240 g within 4 consecutive days and analgesia was assessed using tail-flick test. Chronic dexamethasone was applied using 4 daily i.p. injections. Lumbar spinal tissues were assayed for the expression of G_αi/o and G_β proteins using “semiquantitative PCR” normalized to beta-actin gene expression.
Results showed that chronic administration of dexamethasone could reduce and reverse the development of tolerance in rats that received chronic i.p. injections of morphine. Chronic administration of dexamethasone significantly increased the expression of G_αi/o, while chronic administration of morphine did not change its expression. The expression of G_β, however, was increased after the chronic administration of morphine, but did not change after the administration of chronic dexamethasone. None of these increases were observed when morphine and dexamethasone were co-administered.
We conclude that the development of tolerance to analgesic effect of morphine could be prevented and reversed by dexamethasone co-administration. The increase in G_αi/o genes expression produced by chronic dexamethasone may facilitate the opioid signaling pathway and compensate for morphine-induced tolerance.
Changes in G proteins genes expression in rat lumbar spinal cord support the inhibitory effect of chronic pain on the development of tolerance to morphine analgesia
2005, Neuroscience Research
There are some reports regarding the inhibitory effect of pain on tolerance development to analgesic effect of opioids. The present study was designed to investigate whether the chronic formalin induced pain is able to reverse analgesic tolerance to morphine and to evaluate the expression of G_αi/o and G_β subunits of G proteins in the context of chronic pain, development of morphine tolerance and their combination.
Morphine tolerance was induced by chronic systemic (intraperitoneally, i.p.) or spinal (intrathecally, i.t.) administration of morphine to male Wistar rats weighing 200–240 g and analgesia was assessed using tail flick test. Chronic pain was induced by 4 daily intraplantar injections of 50 μl of 5% formalin. Lumbar spinal tissues were assayed for the expression of G_αi/o and G_β proteins using “semiquantitative PCR” normalized to beta-actin gene expression.
Results showed that chronic formalin induced pain could reduce and reverse the development of tolerance in rats that had received chronic (i.p. or i.t.) administration of morphine. Chronic administration of morphine did not change G_αi/o gene expression, while chronic pain significantly increased its expression. The expression of G_β, however, was increased after the chronic administration of morphine, but did not change after the induction of chronic pain. None of these increases were observed when morphine and formalin were administered at the same time.
Due to synchronous development of morphine tolerance and changes in expression of G_β, it may be concluded that the development of tolerance to analgesic effect of morphine is partially mediated by increase in G_β gene expression. The increase in G_αi/o genes expression produced by chronic pain may facilitate the opioid signaling pathway and compensate for morphine-induced tolerance.
μ opioid receptor signaling in morphine sensitization
2003, Neuroscience
We used a previously reported model of morphine sensitization that elicited a complex behavioral syndrome involving stereotyped and non stereotyped activity. To identify the mechanism of these long-lasting processes, we checked the density of μ opioid receptors, receptor-G-protein coupling and the cyclic AMP (cAMP) cascade. In morphine-sensitized animals μ opioid receptor autoradiography revealed a significant increase in the caudate putamen (30% versus controls), nucleus accumbens shell (16%), prefrontal and frontal cortex (26%), medial thalamus (43%), hypothalamus (200%) and central gray (89%). Concerning morphine’s activation of G proteins in the brain, investigated in the guanylyl 5′-[γ-³⁵S]thio]triphosphate ([³⁵S]GTPγS) binding assay, a significant increase in net [³⁵S]GTPγS binding was seen in the caudate putamen (39%) and hypothalamus (27%). In the caudate putamen this was due to an increase in the amount of activated G proteins, and in the hypothalamus to a greater affinity of G proteins for guanosine triphosphate (GTP). The main second messenger system linked to the opioid receptor is the cAMP pathway. In the striatum basal cAMP levels were significantly elevated in sensitized animals (70% versus controls) and [D-Ala², N-Me-Phe⁴, Gly⁵-ol]-enkephalin (DAMGO) significantly inhibited forskolin-stimulated cAMP production in control (30%) but not in sensitized rats. In the hypothalamus no significant changes were observed in basal cAMP levels and DAMGO inhibition.
These cellular events induced by morphine pre-exposure could underlie the neuroadaptive processes involved in morphine sensitization.

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Regular paperAdaptive changes in the number of Gs- and Gi-proteins underlie adenylyl cyclase sensitization in morphine-treated rat striatal neurons

Abstract

Eur. J. Pharmacol. Mol. Pharmacol.

Trends Neurosci.

Trends Pharmacol. Sci.

Trends Pharmacol. Sci.

Life Sci.

J. Biol. Chem.

Anal. Biochem.

Eur. J. Pharmacol.

Brain Res.

Trends Pharmacol. Sci.

Life Sci.

Eur. J. Pharmacol. Mol. Pharmacol.

Eur. J. Pharmacol. Mol. Pharmacol.

Stoichiometry of receptor-Gs-adenylate cyclase interactions

FASEB J.

Use of antisense to study the role of Gs alpha subunit in catecholamine stimulation to adenylate cyclase

Long-term opiate exposure leads to reduction of the αi-1 subunit of GTP-binding proteins

J. Neurochem.

A primary culture of striatal neurons: a model of choice for pharmacological and biochemical studies of neurotransmitter receptors

J. Physiol. (Paris)

Regular paper
Adaptive changes in the number of G_s- and G_i-proteins underlie adenylyl cyclase sensitization in morphine-treated rat striatal neurons

Stoichiometry of receptor-G_s-adenylate cyclase interactions

Use of antisense to study the role of G_s alpha subunit in catecholamine stimulation to adenylate cyclase