h-89 and Substance-Withdrawal-Syndrome

The present study shows interactive effects of bucladesine (db-cAMP) as a cyclic adenosine monophosphate (cAMP) agonist and H-89 as a protein kinase A (PKA) inhibitor on naloxone-induced withdrawal signs in morphine-dependent mice. Animals were treated subcutaneously with morphine thrice daily with doses progressively increased from 50 to 125 mg/kg. A last dose of morphine (50 mg/kg) was administered on the 4th day. Several withdrawal signs were precipitated by intraperitoneal (i.p.) administration of naloxone (5 mg/kg). Different doses of bucladesine (50, 100, 200 nm/mouse) and H-89 (0.05, 0.5, 1, 5 mg/kg) were administered (i.p.) 60 min before naloxone injection. In combination groups, bucladesine was injected 15 min before H-89 injection. Single administration of H-89 (0.5, 1, 5 mg/kg) and bucladesine (50, 100 nm/mouse) significantly attenuated prominent behavioral signs of morphine withdrawal. Lower doses of bucladesine (50, 100 nm/mouse) in combination with H-89 (0.05 mg/kg) increased the inhibitory effects of H-89 on withdrawal signs while in high dose (200 nm/mouse) decreased the ameliorative function of H-89 (0.05 mg/kg) in morphine-dependent animals. It is concluded that H-89 and bucladesine could affect morphine withdrawal syndrome via possible interaction with cyclic nucleotide messengering systems, protein kinase A signaling pathways, and modified related neurotransmitters.

Topics: Animals; Bucladesine; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Dose-Response Relationship, Drug; Drug Therapy, Combination; Isoquinolines; Male; Mice; Morphine Dependence; Naloxone; Narcotic Antagonists; Substance Withdrawal Syndrome; Sulfonamides

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 A(1) 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 micro-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 micro-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 micro-receptor coupling to presynaptic inhibition that overcomes an underlying tolerance.

Topics: Action Potentials; Adenosine; Affinity Labels; Animals; Colforsin; Cyclic AMP; Dipyridamole; Dose-Response Relationship, Drug; Drug Interactions; Enkephalins; Enzyme Inhibitors; gamma-Aminobutyric Acid; In Vitro Techniques; Isoquinolines; Male; Mesencephalon; Mice; Mice, Inbred C57BL; Morphine; Morphine Dependence; Naloxone; Narcotic Antagonists; Narcotics; Neural Inhibition; Neurons; Patch-Clamp Techniques; Periaqueductal Gray; Probenecid; Purinergic P1 Receptor Agonists; Purinergic P1 Receptor Antagonists; Substance Withdrawal Syndrome; Sulfonamides; Synaptic Transmission; Thioinosine; Time Factors; Uricosuric Agents; Vasodilator Agents; Xanthines

Major adaptations after chronic exposure to morphine include an up-regulation of the adenosine 3',5'-monophosphate pathway. Acute opioids, via mu-opioid receptors, disinhibit midbrain serotonergic neurons by suppressing inhibitory GABAergic transmission in the dorsal raphe nucleus and adjacent periaqueductal gray. This study examined whether chronic morphine induces a compensatory increase in GABA inputs to 5-hydroxytryptamine neurons and whether this was associated with an up-regulation of the adenosine 3',5'-monophosphate pathway. The firing rate of serotonergic neurons was reduced in brain slices from morphine-dependent rats, an effect reversed by the GABA(A) antagonist bicuculline. The reduction in firing rate was accompanied by an increased frequency of spontaneous GABAergic inhibitory postsynaptic currents, indicating increased GABA tone in the slice. The increase in GABA tone in brain slices from dependent rats was associated with increased induction of inhibitory postsynaptic currents by the adenylyl cyclase activator forskolin, suggesting an up-regulation of the adenosine 3',5'-monophosphate pathway. Indeed, chronic morphine increased levels of adenylyl cyclase VIII (but not of adenylyl cyclase I, III or V) immunoreactivity in the dorsal raphe nucleus area. Two adenosine 3',5'-monophosphate-mediated mechanisms for the increase in GABA tone were discerned. The first, which predominated when impulse-flow was blocked by tetrodotoxin, involves protein kinase A since it was sensitive to protein kinase A inhibitors. The second, seen when impulse-flow was intact (i.e. absence of tetrodotoxin), was insensitive to protein kinase A inhibitors but was suppressed by ZD7288, a blocker of hyperpolarizing-activated Ih channels which are directly activated by adenosine 3',5'-monophosphate. We conclude that chronic morphine induces an up-regulation of the adenosine 3',5'-monophosphate pathway in GABAergic inputs to serotonergic cells, resulting in an increase in spontaneous and impulse-flow dependent GABA release. These changes would lead to an increase in GABA tone and subsequently to the reported decrease in serotonergic activity during opiate withdrawal.

Topics: Adenylyl Cyclases; Analgesics, Opioid; Animals; Cardiovascular Agents; Colforsin; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Electrophysiology; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enzyme Inhibitors; Excitatory Amino Acid Agonists; gamma-Aminobutyric Acid; In Vitro Techniques; Interneurons; Isoquinolines; Male; Membrane Potentials; Morphine; N-Methylaspartate; Neural Inhibition; Periaqueductal Gray; Phenylephrine; Pyrimidines; Raphe Nuclei; Rats; Rats, Sprague-Dawley; Serotonin; Substance Withdrawal Syndrome; Sulfonamides; Sympathomimetics; Tetrazoles; Tetrodotoxin; Thionucleotides