title: Prolonged activation of Ca2+-activated K+ current contributes to the long-lasting refractory period of Aplysia bag cell neurons.
abstract: Stimulation of the bag cell neurons of Aplysia activates several biochemical pathways, including protein kinase C (PKC), and alters their excitability for many hours. After an approximately 30 min afterdischarge, these neurons enter an approximately 18 hr inhibited state during which additional stimulation fails to evoke discharges. In vivo, this refractory period limits the frequency of reproductive behaviors associated with egg laying. We have now examined the role of Ca2+-activated K+ (BK) currents in the refractory period. Outward currents gated by both intracellular Ca2+ and depolarization, with pharmacological characteristics of BK currents, were recorded in isolated bag cell neurons. These currents were enhanced by the BK channel activators phloretin and 1,3-dihydro-1-[2-hydroxy-5-(trifluoro-methyl)phenyl]-5-trifluoromethyl-2H-benzimidazol-2-one and inhibited by the BK blocker paxilline. The BK component of K+ current was enhanced by 12-O-tetradecanoyl-phorbol-13-acetate, an activator of PKC, and this effect was blocked by sphinganine and PKC(19-36), inhibitors of PKC in bag cell neurons. To test whether the BK current is altered during the refractory period, intact clusters were stimulated to afterdischarge, and neurons were isolated after the clusters had entered the refractory period. Compared with unstimulated cells, current density was almost doubled in refractory neurons. This increase in current was inhibited by preincubating clusters in sphinganine. Treatment of refractory clusters with paxilline significantly restored the ability of stimulation to evoke afterdischarges. Conversely, application of phloretin to previously unstimulated clusters inhibited the onset of afterdischarges. These results indicate that a prolonged increase in BK channel activity contributes to the prolonged refractory period of the bag cell neurons.
authors.affiliation: Department of Pharmacology, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
authors.forename: Yalan
authors.initials: Y
authors.lastname: Zhang
authors.forename: Neil S
authors.initials: NS
authors.lastname: Magoski
authors.forename: Leonard K
authors.initials: LK
authors.lastname: Kaczmarek
mesh_terms: D000818:Animals; D001048:Aplysia; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D002478:Cells, Cultured; D004558:Electric Stimulation; D020536:Enzyme Activators / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D051036:Large-Conductance Calcium-Activated Potassium Channels; D009433:Neural Inhibition / Q000187:drug effects / Q000502:physiology*; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D010693:Phloretin / Q000494:pharmacology; D011188:Potassium / Q000378:metabolism; D026902:Potassium Channel Blockers / Q000494:pharmacology; D024681:Potassium Channels, Calcium-Activated / Q000187:drug effects / Q000378:metabolism*; D011493:Protein Kinase C / Q000187:drug effects / Q000378:metabolism; D012032:Refractory Period, Electrophysiological / Q000187:drug effects / Q000502:physiology*; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
chemical_list: D020536:Enzyme Activators; D004791:Enzyme Inhibitors; D051036:Large-Conductance Calcium-Activated Potassium Channels; D026902:Potassium Channel Blockers; D024681:Potassium Channels, Calcium-Activated; D011493:Protein Kinase C; D013110:Sphingosine; D013755:Tetradecanoylphorbol Acetate; C005682:safingol; D011188:Potassium; D010693:Phloretin; D002118:Calcium