PURINERGIC MODULATION OF ADULT GUINEA-PIG CARDIOMYOCYTES IN LONG-TERMCULTURES AND COCULTURES WITH EXTRACARDIAC OR INTRINSIC CARDIAC NEURONS

Objective: To determine the capacity of ATP to modify cardiomyocytes d irectly or indirectly via peripheral autonomic neurones, the effects o f various purinergic agents were studied on long term cultures of adul t guinea pig ventricular myocytes and their co-cultures with extracard iac (stellate ganglion) or intrinsic cardiac neurones. Methods: Ventri cular myocytes and cardiac neurones were enzymatically dissociated and plated together or alone (myocytes only). Myocyte cultures were used for experiments after three to six weeks. The electrical and contracti le properties of cultured myocytes and myocyte-neuronal networks were investigated. Results: The spontaneous beating frequency of ventricula r myocytes co-cultured with stellate ganglion neurones increased by si milar to 140% (p<0.001) following superfusion with 10(-5) M ATP. This effect was not modified significantly by tetrodotoxin or by beta adren oceptor blockade (10(-5) M timolol), but was eliminated following appl ication of the P-2 antagonist suramin (10(-5) M). Basal spontaneous co ntractile rate was reduced by similar to 86% (p<0.001) in the presence of suramin, indicating the existence of tonically active purinergic s ynaptic mechanisms in stellate ganglion neurone-myocyte cocultures. Su ramin did not significantly affect non-innervated myocyte cultures. AT P increased myocyte contractile rate in intrinsic cardiac neurone-myoc yte co-cultures by similar to 40% (p<0.01) under control conditions, b ut when beta adrenergic receptors of tetrodotoxin sensitive neural res ponses were blocked, ATP induced greater augmentation (>100%). In cont rast, ATP induced much smaller effects in non-innervated myocyte cultu res (similar to 26%, p<0.01). Analogues of ATP showed the following or der of potency: ATP > UTP > MSATP > beta gamma ATP > alpha beta ATP. A denosine (10(-4) M) attenuated the beating frequency of myocytes in bo th types of co-culture, while not significantly affecting non-innervat ed myocyte cultures. Conclusions: The experimental model used in this study showed that extrinsic and intrinsic cardiac neurones which posse ss P-2 receptors can greatly enhance cardiac myocyte contractile rate when activated by ATP. Since adenosine reduced contractile rate in bot h types of co-cultures while not affecting non-innervated myocytes, it is concluded that some of these neurones possess P-1 receptors.