Reports have suggested that the fast inward sodium current (INa) in cardiac tissues may be modulated by β-adrenergic stimulation and that such modulation may affect conduction in the setting of myocardial ischemia and infarction. However, many of these studies have used dissociated myocytes or broken cell preparations, whose responses need not necessarily reflect those of syncytial preparations. To investigate further the possibility that β-adrenergic stimulation of INamay differ in various preparations, we compared the effects of the β-agonist isoproterenol (ISO) on syncytial canine Purkinje fibers and ventricular muscle preparations, as well as isolated ventricular myocytes. Alterations of the maximum rate of rise of the action potential upstroke (Vmax) were used as an index of changes of INa. ISO (1 μM) had no effect on Vmaxof upstrokes of normally polarized (fast responses) or partially depolarized (elevated [K+]0, depressed fast responses) syncytial ventricular muscle preparations or Purkinje fibers. In contrast, lower concentrations of ISO (0.5–1.0 μM) modulated Vmaxof isolated ventricular myocytes, depending on the technique used to monitor transmembrane potential. When 2.7 M KCI-filled microelectrodes were used, ISO reduced Vmaxof partially depolarized myocytes without affecting Vmaxof normally polarized myocytes. However, when myocytes were dialyzed using patch pipettes, ISO reduced Vmaxof partially depolarized myocytes and increased Vmaxof normally polarized myocytes, effecting a hyperpolarized shift of the normalized inactivation curve relating Vmaxto resting membrane potential. The different β-adrenergic responses of syncytial preparations and nondialyzed and dialyzed myocytes suggest that differences in the ionic or metabolic condition of the preparations likely alter cAMP-dependent responses and channel phosphorylation. These results suggest that β-adrenergic modulation of INacan occur under some experimental conditions but that extrapolation of data obtained using isolated myocytes to syncytial preparations in vitro or in vivo requires further evaluation.