Determinants of atrial natriuretic peptide secretion in cultured atrial myocytes

Atrial natriuretic peptide (ANP) secretion by atrial myocytes in 7- to 8-day primary cultures from adult rats was measured by radioimmunoassay under conditions designed to separate primary effects on secretion from effects caused by contractions. Abolishing contraction with 10 μM tetrodotoxin significantly reduced ANP accumulated in 2 h. Raising external Ca²⁺ concentration from 0.2 to 1.2 mM in the presence of tetrodotoxin did not increase ANP secretion. Substantial ANP secretion persisted in a nominally Ca²⁺-free medium containing 10 mM ethylene glycol-bis(β-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) and was not diminished by 100 μM ryanodine. In the presence of EGTA, 100 nM 12-O-tetradecanoylphorbol 13-acetate (TPA) significantly increased ANP secretion; this increment was unaffected by 100 μM ryanodine. These experiments suggest 1) that in absence of contractions, ANP secretion requires neither transplasmalemmal Ca²⁺ influx nor ryanodine-inhibitable Ca²⁺ release by sarcoplasmic reticulum (SR) and 2) that TPA stimulates ANP secretion without requiring Ca influx or ryanodine-inhibitable SR Ca release.