Detection and photoaffinity labeling of the Ca2+-activated K+ channel-associated apamin receptor in cultured astrocytes from rat brain.
Apamin, an 18-amino acid bee venom peptide, is a specific blocker of a class of Ca2+ activated K+ channels. Mono 125I-iodoapamin was used to detect the K+ channel-associated receptor site in cultured astrocytes from rat brain. Specific high-affinity binding to intact glial cells with a Kd of about 90 pM at 1 degree C and pH 7.5 was demonstrated by equilibrium and kinetic methods. The average receptor capacity was 3 fmol/mg cell protein which is 2 to 3-fold lower than in primary cultured neurons. Binding was stimulated by K+ ions, but to a lesser extent than with neuronal receptors. Photoaffinity labeling of receptor/ion channel components using an arylazide derivative of 125I-monoiodoapamin revealed the presence of the 86- and 33-kDa polypeptides, previously detected in neurones. However a 59-kDa peptide which is present in synaptic membrane preparations from adult rat brain, but not in cultured neurons, was also clearly labeled in intact astrocytes. This indicates that the 59-kDa polypeptide is not a proteolytic fragment of the 86-kDa chain but an associated subunit which is only accessible to photolabeling in certain apamin receptor preparations. Apamin-sensitive Ca2+-activated K+ channels in astrocytes may be one of the pathways by which glial cells redistribute K+ in the central nervous system (CNS).
D000345:Affinity Labels; D000818:Animals; D001030:Apamin / Q000378:metabolism; D001253:Astrocytes / Q000378:metabolism*; D001921:Brain / Q000166:cytology / Q000378:metabolism*; D002118:Calcium / Q000502:physiology; D002478:Cells, Cultured; D007473:Ion Channels / Q000378:metabolism*; D007700:Kinetics; D008970:Molecular Weight; D011188:Potassium / Q000378:metabolism / Q000502:physiology*; D015221:Potassium Channels; D051381:Rats; D017981:Receptors, Neurotransmitter / Q000378:metabolism*
D000345:Affinity Labels; D007473:Ion Channels; D015221:Potassium Channels; D017981:Receptors, Neurotransmitter; C033799:apamin receptor; D001030:Apamin; D011188:Potassium; D002118:Calcium