Chronic ethanol exposure enhances signaling through muscarinic receptors expressed by cRNA injection in Xenopus oocytes: implications for mechanism of action.
The molecular mechanisms underlying the cerebral symptoms of ethanol withdrawal syndrome are poorly understood. In addition to ethanol's effect on GABA and NMDA receptors, ethanol affects muscarinic acetylcholine signaling. This interaction has attracted attention because of the importance of muscarinic signaling in consciousness. Chronic ethanol exposure increases muscarinic receptor binding. Increased transcription of receptor message has been suggested as the underlying mechanism, but this hypothesis has not been tested directly. Therefore, we studied the effects of ethanol on muscarinic signaling in a model that bypasses transcription of muscarinic receptor genes. We expressed rat m1 muscarinic receptors by cRNA microinjection in Xenopus oocytes. Cells were voltage-clamped at -70 mV and effects of prolonged (24, 48, and 72 hr) exposure to ethanol (25, 50, and 100 mM) on methylcholine-induced calcium-activated Cl- currents were determined. Effects of prolonged ethanol exposure on currents induced by stimulation of lysophosphatidate receptors, direct G protein activation, or inositol trisphosphate receptor activation were studied as well. Prolonged ethanol exposure enhanced methylcholine (or lysophosphatidate-)-induced currents in a time- and concentration-dependent manner. Thus, enhanced muscarinic gene transcription is not required for ethanol enhancement of muscarinic signaling. Lack of ethanol effect on inositol trisphosphate-induced signaling suggests that intracellular signaling systems downstream of phospholipase C are not involved. In contrast, currents induced by direct G protein stimulation were enhanced significantly. Therefore, one potential site of ethanol's action on muscarinic signaling is upregulation of the associated G protein or enhancement of its functioning.
Hönemann
C W
CW
Department of Anesthesiology, University of Virginia, Charlottesville 22906-0010, USA.
Wong
A
A
Arledge
J A
JA
Durieux
M E
ME
D000818:Animals; D000431:Ethanol / Q000494:pharmacology*; D019204:GTP-Binding Proteins / Q000187:drug effects / Q000502:physiology; D008845:Microinjections; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism*; D018075:RNA, Complementary / Q000008:administration & dosage / Q000494:pharmacology; D051381:Rats; D011976:Receptors, Muscarinic / Q000187:drug effects* / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D014981:Xenopus / Q000378:metabolism* / Q000502:physiology
D018075:RNA, Complementary; D011976:Receptors, Muscarinic; D000431:Ethanol; D019204:GTP-Binding Proteins