Adrenergic stimulation of substrate utilization by cardiac myocytes isolated from rainbow trout

A method is described for the isolation of calcium-tolerant myocytes from adult rainbow trout. Isolated myocytes remain viable for at least 4 h in suspension as indicated by (1) maintenance of ATP, phosphocreatine (PCr) and glycogen levels; (2) maintenance of the integrity of cell membranes, shown by low rates of leakage of lactate dehydrogenase (LDH) to the medium and exclusion of Trypan Blue; (3) the ability to metabolize substrates; and (4) sensitivity to adrenergic agonists. CO2 production from both glucose and lactate was sensitive to adrenergic stimulation, with the following order of potency: isoproterenol greater than noradrenaline much greater than adrenaline greater than phenylephrine, which indicates the presence of beta 1-adrenoceptors. Myocytes isolated from trout acclimated to 20 degrees C in the summer were more sensitive to beta-adrenergic stimulation than myocytes isolated from trout acclimated to 9 degrees C in either summer or winter. In the absence of exogenous fuel, there was a net reduction in myocyte glycogen content and glycogenolysis was further stimulated by 10(-7) mmol l-1 noradrenaline. However, in the presence of exogenous fuel (either 5 mmol l-1 lactate or 5 mmol l-1 glucose), glycogen was 'spared' and noradrenaline-stimulated glycogenolysis was apparently inhibited.