A major focus of our work involves an evaluation of the acute and chronic effects of ethanol in the CNS. However, the brain represents a heterogeneous collection of cell types, and distinction of direct and indirect effects of ethanol can be difficult. We are developing in vitro organ and tissue culture systems that can be used to monitor specific, direct effects of ethanol, for comparison and contrast with results obtained in brain tissue and in vivo. We have developed a primary culture of cerebellar granule cells in which beta-adrenergic receptors and agonist-stimulated cyclic AMP production can be measured; the role of glial and neuronal elements in these responses is being analyzed. Stimulation of cyclic GMP production in cerebellar granule cells is also being determined. Ethanol appears to inhibit this stimulation, similar to results reported in brain. Ethanol enhances isoproterenol-stimulated cyclic AMP and melatonin production by pineal glands in culture. These findings of enhanced receptor-effector coupling are similar to what we have reported in certain brain areas, and allow and investigation of the effects of ethanol on the physiological consequences of receptor activation. Ethanol also appears to enhance cyclic AMP production in pheochromocytoma (PC12) cells, and these cells are being characterized with respect to stimulation of cyclic AMP production by vasoative intestinal peptide, adenosine and forskolin. In PC12 cells, we have also found that low concentrations of ethanol (25mM) inhibit muscarine- and depolarization-induced norepinephrine release. In the former case, ethanol appears to modulate muscarine-stimulated inositol trisphosphate production, and therefore calcium mobilization. These systems allow for detailed examination of the sites and mechanisms of action of ethanol and of changes in biochemical systems that may, in vivo, be associated with ethanol tolerance and physical dependence.
