Arachidonic acid stimulates protein kinase C-epsilon redistribution in heart cells.

Arachidonic acid is elevated in a variety of cell types in response to extracellular stimuli, and has been hypothesized to exert at least some of its intracellular actions via activation of protein kinase C. Here we show that arachidonic acid stimulates a unique pattern of translocation of the epsilon-isoform of protein kinase C in isolated adult rat cardiac myocytes. Using western blot analysis, the majority of epsilon-protein kinase C was found in a cytosolic fraction in unstimulated cells. Treatment with 50 microM arachidonic acid caused a transient increase of epsilon-protein kinase C in a membrane fraction within 1 minute, then after 5-20 minutes most was found in a filament/nuclear fraction. Immunofluorescence and confocal microscopy of the filament fraction revealed a striated staining pattern with epsilon-protein kinase C localized near the Z-line where actin filaments are anchored and where transverse tubules are closely apposed to the myofilaments. delta-Protein kinase C, another isoform highly expressed in these cells, did not redistribute significantly in response to arachidonic acid, but in response to phorbol ester displayed a predominantly nuclear localization. Arachidonic acid also stimulated phosphorylation of the thin filament protein, troponin I, consistent with a filament localization for activated PKC. The physiological relevance of these findings was supported by the observation that 50 microM arachidonic acid promoted a 2.3-fold enhancement of myocyte twitch amplitude, an effect that was significantly blocked by the protein kinase C antagonist chelerythrine. Moreover, the onset of this physiological response correlated in time with translocation of epsilon-protein kinase C to the filaments. The results suggest that arachidonic acid initiates a redistribution of epsilon-protein kinase C to myofilament structures at or near the Z-line where this isozyme would be strategically located to regulate myofilament function and excitation-contraction coupling.