Molecular and functional identification of cyclic AMP-sensitive BKCa potassium channels (ZERO variant) and L-type voltage-dependent calcium channels in single rat juxtaglomerular cells.
This study aimed at identifying the type and functional significance of potassium channels and voltage-dependent calcium channels (Ca(v)) in single rat JG cells using whole-cell patch clamp. Single JG cells displayed outward rectification at positive membrane potentials and limited net currents between -60 and -10 mV. Blockade of K+ channels with TEA inhibited 83% of the current at +105 mV. Inhibition of KV channels with 4-AP inhibited 21% of the current. Blockade of calcium-sensitive voltage-gated K+ channels (BKCa) with charybdotoxin or iberiotoxin inhibited 89% and 82% of the current, respectively. Double immunofluorescence confirmed the presence of BKCa and renin in the same cell. cAMP increased the outward current by 1.6-fold, and this was inhibited by 74% with iberiotoxin. Expression of the cAMP-sensitive splice variant (ZERO) of BKCa was confirmed in single-sampled JG cells by RT-PCR. The resting membrane potential of JG cells was -32 mV and activation of BKCa with cAMP hyperpolarized cells on average 16 mV, and inhibition with TEA depolarized cells by 17 mV. The cells displayed typical high-voltage activated calcium currents sensitive to the L-type Ca(v) blocker calciseptine. RT-PCR analysis and double-immunofluorescence labeling showed coexpression of renin and L-type Ca(v) 1.2. The cAMP-mediated increase in exocytosis (measured as membrane capacitance) was inhibited by depolarization to +10 mV, and this inhibitory effect was blocked with calciseptine, whereas K+-blockers had no effect. We conclude that JG cells express functional cAMP-sensitive BKCa channels (the ZERO splice variant) and voltage-dependent L-type Ca2+ channels.
Friis
Ulla G
UG
Department of Physiology and Pharmacology, University of Southern Denmark, Denmark. ufriis@health.sdu.dk
Jørgensen
Finn
F
Andreasen
Ditte
D
Jensen
Boye L
BL
Skøtt
Ole
O
D017398:Alternative Splicing; D000818:Animals; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D020746:Calcium Channels, L-Type / Q000096:biosynthesis / Q000187:drug effects / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism / Q000502:physiology; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism* / Q000494:pharmacology; D025802:Electric Capacitance; D015640:Ion Channel Gating / Q000187:drug effects; D007606:Juxtaglomerular Apparatus / Q000166:cytology / Q000378:metabolism*; D051036:Large-Conductance Calcium-Activated Potassium Channels; D008297:Male; D018408:Patch-Clamp Techniques; D011188:Potassium / Q000378:metabolism; D026902:Potassium Channel Blockers / Q000494:pharmacology; D024681:Potassium Channels, Calcium-Activated / Q000096:biosynthesis / Q000145:classification / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012083:Renin / Q000378:metabolism; D012964:Sodium / Q000378:metabolism
D020746:Calcium Channels, L-Type; C419385:L-type calcium channel alpha(1C); D051036:Large-Conductance Calcium-Activated Potassium Channels; D026902:Potassium Channel Blockers; D024681:Potassium Channels, Calcium-Activated; D012964:Sodium; D000242:Cyclic AMP; D012083:Renin; D011188:Potassium; D002118:Calcium