Ca2 Transients and Ca2 Waves in Purkinje Cells: Role in Action Potential Initiation

ABSTRACTPurkinje cells contain sarcoplasmic reticulum (SR) directly under the surface membrane, are devoid of t-tubuli, and are packed with myofibrils surrounded by central SR. Several studies have reported that electrical excitation induces a biphasic Ca transient in Purkinje fiber bundles. We determined the nature of the biphasic Ca transient in aggregates of Purkinje cells. Aggregates (n=12) were dispersed from the subendocardial Purkinje fiber network of normal canine left ventricle, loaded with Fluo-3/AM, and studied in normal Tyrode’s solution (24°C). Membrane action potentials were recorded with fine-tipped microelectrodes, and spatial and temporal changes in [Ca]i were obtained from fluorescent images with an epifluorescent microscope (×20; Nikon). Electrical stimulation elicited an action potential as well as a sudden increase in fluorescence (L0) compared with resting levels. This was followed by a further increase in fluorescence (L1) along the edges of the cells. Fluorescence then progressed toward the Purkinje cell core (velocity of propagation 180 to 313 μm/s). In 62% of the aggregates, initial fluorescent changes of L0 were followed by focally arising Ca waves (L2), which propagated at 158±14 μm/s (n=13). Spontaneous Ca waves (L2*) propagated like L2 (164±10 μm/s) occurred between stimuli and caused slow membrane depolarization; 28% of L2* elicited action potentials. Both spontaneous Ca wave propagation and resulting membrane depolarization were thapsigargin sensitive. Early afterdepolarizations were not accompanied by Ca waves. Action potentials in Purkinje aggregates induced a rapid rise of Ca through ICaL and release from a subsarcolemmal compartment (L0). Ca release during L0 either induced further Ca release, which propagated toward the cell core (L1), or initiated Ca release from small regions and caused L2 Ca waves, which propagated throughout the aggregate. Spontaneous Ca waves (L2*) induce action potentials.