The effects of tetrodotoxin on sinus node activity were studied by direct perfusion through its nutrient artery. Tetrodotoxin suppresses action potentials by blocking sodium entry into the cell. In the open-chest anesthetized dog, direct perfusion of the sinus node artery with tetrodotoxin, 1.0 μg/ml, caused immediate sinus slowing lasting 12 to 60 minutes. This slowing was not affected by intranodal atropinization. Tetrodotoxin also produced complete blockade of sinus node response to vagal stimulation. Blockade of sinus node responses to stellate stimulation was less profound. Tetrodotoxin had no effect on chronotropic responses to intranodal acetylcholine and norepinephrine. These findings suggest that blockade of transmembrane sodium transport (or the rapid entry channel) has a negative chronotropic effect which is not cholinergic and during which response to neurotransmitter substances is normal. Concomitant with this effect there is blockade of neurotransmitter release (or synthesis) in the sinus node.