Early effects of PRL on ion conductances in CHO cells expressing PRL receptor.
Chinese hamster ovary (CHO-K1) cells were stably transfected with prolactin (PRL) receptor cDNA. These cells (CHO-E32) expressed the long form of functional PRL receptor. Using microfluorimetric and patch-clamp techniques, we have investigated the effects of PRL on intracellular Ca2+ concentration ([Ca2+]i) and membrane ion conductances. Exposure of CHO-E32 cells to 5 nM PRL resulted in an increase in [Ca2+]i. Two types of response were observed: 1) a stimulation of Ca2+ entry and 2) an intracellular Ca2+ mobilization. As PRL inhibited voltage-activated Ca2+ current, the PRL-induced Ca2+ increase does not involve voltage-activated Ca2+ channels. PRL also increased a charybdotoxin-sensitive Ca(2+)-dependent K+ conductance. Simultaneous measurements showed that PRL hyperpolarized the membrane potential before increasing intracellular Ca2+ levels. In voltage clamp, hyperpolarizing voltage steps were associated with increased Ca2+ concentrations, whereas depolarizing voltage steps decreased [Ca2+]i. Cell-free patch-clamp experiments showed that PRL directly stimulates K+ channel activity. Our results suggest the existence of a regulatory complex involving a protein kinase tightly associated with the Ca(2+)-activated K+ channels and that PRL stimulates these channels by means of the activation of protein kinase. The resulting hyperpolarization stimulates Ca2+ entry, probably through voltage-insensitive nonspecific channels.
Prevarskaya
N
N
Laboratory of Neurophysiology, University of Bordeaux II, France.
Skryma
R
R
Vacher
P
P
Daniel
N
N
Bignon
C
C
Djiane
J
J
Dufy
B
B
D000818:Animals; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism / Q000502:physiology*; D002474:Cell-Free System; D018999:Charybdotoxin; D006224:Cricetinae; D018076:DNA, Complementary; D004553:Electric Conductivity; D005470:Fluorometry; D007425:Intracellular Membranes / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects; D011188:Potassium / Q000502:physiology*; D011388:Prolactin / Q000494:pharmacology*; D011981:Receptors, Prolactin / Q000235:genetics / Q000378:metabolism*; D012604:Scorpion Venoms / Q000494:pharmacology; D013779:Tetrodotoxin / Q000494:pharmacology; D014162:Transfection
D018076:DNA, Complementary; D011981:Receptors, Prolactin; D012604:Scorpion Venoms; D018999:Charybdotoxin; D013779:Tetrodotoxin; D011388:Prolactin; D011188:Potassium; D002118:Calcium