Оbestatin is a gastrointestinal hormone with various peripheral and central effects. It induces a positive inotropic effect on excised frog heart preparation, which consists of cardiac muscle tissue with preserved endocardium and functionally active projections of autonomic neurons. The hypothesis that obestatin targets sympathetic nerve endings was checked. The inhibitor of vesicular monoamine transporter reserpine (10 µM), norepinephrine reuptake inhibitor desipramine (1 µM) and tyrosine hydroxylase inhibitor 3-Iodothyrosine (3-IT; 0.02 mg/g b.w.) were administered prior to obestatin application to study the involvement of each of these molecules in the observed physiological response. The proper adrenergic signaling was tested at the end of experiments with 50 µM epinephrine. The positive inotropic effect of obestatin was abolished in the presence of 3-IT and reserpine, while desipramine just slightly reduced the observed effect (by up to 50%). It is concluded that the positive inotropic effect of obestatin is mediated by a neuronal-dependent mechanism. It is suggested that this regulation is probably realized mainly by an increase in epinephrine secretion.