Characterization of α1-adrenoceptors which mediate chronotropy in neonatal rat cardiac myocytes

Abstract

1. In the present study, we investigated the effect of culture on α₁-adrenoceptors that mediate chronotropy and on α₁-adrenergic signal transduction in neonatal rat cardiac myocytes.

2. The spontaneous beating rate of neonatal rat myocytes after 3 or 7 days in culture was 37.4 ± 4.2 or 102.0 ± 4.3 beats min⁻, respectively. The α₁-adrenoceptor-mediated chronotropic effect of norepinephrine was positive at day 3 of culture. In contrast to day 3 of culture, the neonatal myocytes exhibited a negative chronotropic response to norepinephrine on day 7 of culture. Both of these effects of norepinephrine were completely abolished by prazosin.

3. The affinity (K_d) and/or density (B_max) of α₁-adrenoceptors labeled with [³H]prazosin in membranes from cultured myocytes were not significantly different between day 3 and day 7 of culture.

4. The expression of G_s, G_i, G_q and G_o, α-subunits in membranes from cultured myocytes was found to be significantly increased with the passage of culture time by immunoblot analysis. In contrast, no significant differences in G_β-subunit expression were observed between day 3 and day 7 of culture.

5. Norepinephrine-stimulated inositol 1,4,5-trisphosphate production by radio-binding protein in neonatal myocytes after 7 days of culture was significantly higher than that of the day 3 counterpart.

6. No significant changes in phospholipid and cholesterol contents in membranes from neonatal myocytes were observed with longer culture times.

7. These results suggest that changes in the responsiveness to α₁-adrenergic stimulation from positive to negative chronotropy during culture of cardiac myocytes are mediated, at least in part, by functional alterations in the α₁-adrenergic signal transduction systems, including both G-protein expression and inositol 1,4,5-trisphosphate production.