Cultured lymphocytes from alcoholic subjects have altered cAMP signal transduction.

Abstract

Previous work has shown that freshly isolated lymphocytes from alcoholic subjects show significantly reduced basal and adenosine receptor-stimulated cAMP levels. This decrease could be due to ethanol-induced cellular adaptation or to a genetic difference in the regulation of cAMP signal transduction. Therefore, we cultured human lymphocytes in defined medium without ethanol for 7-8 days and then examined differences in receptor-dependent cAMP accumulation between lymphocytes from alcoholic and nonalcoholic subjects. After four to six generations in culture without ethanol, lymphocytes from alcoholic subjects have significantly higher cAMP levels than do cells from nonalcoholic subjects. Thus, a difference in cAMP signal transduction is demonstrable in cells from alcoholic subjects grown without ethanol. We also found that cultured lymphocytes from both alcoholic and nonalcoholic subjects show a decrease in receptor-stimulated cAMP levels after exposure to 200 mM ethanol for 48 hr. To determine whether alcoholic subjects have increased sensitivity to ethanol, lymphocytes were exposed to only 100 mM ethanol for 24 hr. Under these conditions, receptor-dependent cAMP levels did not change in cells from nonalcoholic subjects. However, lymphocytes from alcoholic subjects showed a 39% decrease (P < 0.003) in adenosine receptor-stimulated cAMP levels. Taken together, the results show that (i) chronic ethanol treatment in culture reproduces the suppression of cAMP levels found in circulating lymphocytes from alcoholic subjects and (i) despite four to six cell divisions in culture without ethanol, lymphocytes from alcoholic subjects exhibit signiicantly increased adenosine receptor-dependent cAMP levels and increased sensitivity to chronic exposure to ethanol. These rmdings suggest that the suppression of cAMP levels observed in freshly isolated lymphocytes from alcoholic subjects results from both a direct effect of chronic exposure to ethanol and a genetic difference leading to altered cAMP signal transduction.Receptor-stimulated cAMP production is altered after acute and chronic exposure to ethanol (1-12). Although ethanol acutely stimulates receptor-dependent cAMP production, chronic exposure of cultured NG108-15 neural cells to ethanol decreases adenosine receptor-stimulated cAMP levels when measured in the absence of ethanol (8, 9). Similar findings have also been described for f3-adrenergic receptor stimulation of cAMP production in S49 mouse lymphoma cells (6) and prostaglandin E1 receptor stimulation in both NIE cells (7) and NG108-15 neural cells (9).We have suggested that decreased receptor-stimulated cAMP levels have pathophysiologic significance in chronic alcoholism (13). Freshly isolated lymphocytes from alcoholic subjects show a 75% reduction in basal and adenosine receptor-stimulated cAMP levels when compared with ageand sex-matched control subjects and patients with nonalcoholic liver disease (13). The results of that study indicated that reduced cAMP levels in lymphocytes could be used to distin...

Differential expression of guanosine triphosphate binding proteins in men at high and low risk for the future development of alcoholism.

“…Similar to the hypothesis proposed by Nagy et al (13), we speculate that enhanced expression of Gsa sets the stage for the cell to initially undergo more pronounced activation of its AC system in the presence of alcohol, but that is then followed by more rapid and profound desensitization (e.g., tolerance). The enhanced expression of Gsa may explain the apparent phenomena of greater initial sensitivity to ethanol but more rapid development of tolerance to ethanol that develops in the offspring of alcoholics compared to nonalcoholics (6,7).…”

“…Although freshly isolated lymphocytes from alcoholic subjects have reduced cAMP levels, after several generations in alcohol-free cultures, lymphocytes from alcoholics appear more sensitive to factors that stimulate cAMP production than lymphocytes from nonalcoholic controls (13). It is likely that the reduced cAMP levels found in freshly isolated lymphocytes (13) and platelets (36) (36,37). In this scenario, membrane AC activity would be low or unchanged depending on the degree of Gsa compensation.…”

“…Stimulated cAMP production in freshly isolated platelets and lymphocytes is reduced in alcoholics compared to nonalcoholic controls (11,12). Moreover, after multiple generations in culture, lymphocytes from alcoholics not only recover their responsiveness but develop enhanced sensitivity to factors that stimulate cAMP (13). Work by Palmour et al.…”

Biological markers for increased risk of alcoholism and for quantitation of alcohol consumption.

“…Similarly, lymphocytes from actively drinking alcoholics showed an attenuated response to the adenosine agonist plus ethanol. When lymphocytes from type II alcoholics were cultured in the absence of ethanol for 7-8 d, the lymphocytes not only recover responsiveness to A2 agonists but become supersensitive (18). They are also more sensitive to inhibition of adenosine-stimulated cAMP production after chronic exposure (24 h) to ethanol.…”

Platelet Adenylyl Cyclase Activity as a Trait Marker of Alcohol Dependence

“…AC is responsible for the formation of the second messenger adenosine 3Ј:5Ј-cyclic monophosphate (cAMP), which in turn acts on a number of target molecules, particularly cAMP-dependent protein kinase, to control such diverse phenomena as metabolism, gene transcription, and memory. AC activity, measured in either platelets (Lex et al, 1993;Saito et al, 1994;Tabakoff et al, 1988) or lymphocytes (Diamond et al, 1987;Nagy et al, 1988;Waltman et al, 1993) of alcohol-dependent subjects, has been shown to be less responsive to agents known to increase enzymatic activity. More recently, low platelet AC activity has been proposed as a trait marker for predisposition to alcohol dependence (Devor et al, 1991;Tabakoff and Hoffman, 1998).…”