Evidence for selective coupling of α1-adrenergic receptors to phospholipase C-β1 in rat neonatal cardiomyocytes

Activation of phospholipase C (PLC) in neonatal rat cardiomyocytes (NCM) generates primarily inositol 1,4,5-trisphosphate (Ins(1,4,5)P-3) in response to rises in intracellular Ca2+, or inositol 1,4-bisphosphate (Ins(1,4)P-2) in response to norepinephrine (NE) (Matkovich, S. J. and Woodcock, E. A. (2000) J. Biol. Chem. 275,10845-10850). To examine the PLC subtype mediating the alpha (1)-adrenergic receptor response, PLC-beta (1) and PLC-beta (3) were overexpressed in NCM using adenoviral infection (Ad-PLC-beta (1) NCM and Ad-PLC-beta (3) NCM, respectively) and PLC responses assessed from [H-3]inositol phosphate (InsP) generation in the presence of 10 mM LiCl. The [H-3]InsP response to NE (100 muM) was enhanced in Ad-PLC-beta (1) NCM relative to cells infected with blank virus (Ad-MX NCM), but was reduced in Ad-PLC-beta (3) NCM. In contrast, the [H-3]InsP response to ATP (100 muM) was not elevated in Ad-PLC-beta (1) NCM and was enhanced rather than diminished in Ad-PLC-beta (3) NCM, showing that effects of the two PLC-beta isoforms were specific for particular receptor types. PLC-delta (1) overexpression selectively reduced NE-induced [H-3]InsP responses, without affecting the ATP stimulation. The reduced NE response was associated with a selective loss of PLC-beta (1), expression in Ad-PLC-delta (1) NCM. alpha (1)-Adrenergic receptor activation caused phosphorylation of PLC-beta (1), but not PLC-beta (3), whereas stimulation by ATP induced phosphorylation of PLC-beta (3), but not PLC-beta (1). Taken together, these studies provide evidence that NE-stimulated InsP generation in NCM is primarily mediated by PLC-beta (1), despite the presence of both PLC-beta (1) and PLC-beta (3) isoforms.