CA2-INHIBITED NONINACTIVATING K+ CHANNELS IN CULTURED RAT HIPPOCAMPALPYRAMIDAL NEURONS()

1. Whole-cell perforated-patch recording from cultured CA1-CA3 pyramid al neurones from neonatal rat hippocampus (20-22 degrees C; [K+](o) = 2.5 mM) revealed two previously recorded non-inactivating (sustained) K+ outward currents: a voltage-independent 'leak' current (I-leak) ope rating at all negative potentials, and, at potentials greater than or equal to -60 mV, a time- and voltage-dependent 'M-current' (I-K(M). Bo th were inhibited bar 1 mM Ba2+ or 10 mu M oxotremorine-M (Oxo-M). In ruptured-patch recording using Ca2+-free pipette solution, I-leak was strongly enhanced, and was inhibited by 1 mM Ba2+ but unaffected by 0. 5 mM 4-aminopyridine (4-AP), 1 mM tetraethylammonium (TEA) or 1-10 nM margatoxin. 2. Single channels underlying these currents were sought i n cell-attached patch recordings. A single class of channels of conduc tance similar to 7 pS showing sustained activity at resting potential and above was identified. These normally had a very low open probabili ty (P-o < 0.1), which, however, showed a dramatic and reversible incre ase (to about 0.9 at similar to 0 mV) following the removal of Ca2+ fr om the bath. Under these (Ca2+-free) conditions, single-channel P-o sh owed both voltage-dependent and voltage-independent components on patc h depolarization from resting potential. The mean activation curve was fitted by a modified Boltzmann equation. When tested, all channels we re reversibly inhibited by addition of 10 mu M Oxo-M to the bath solut ion. 3. The channels maintained their high P-o in patches excised in i nside-out mode into a Ca2+-free internal solution and were strongly in hibited by application of Ca2+ to the inner face of the membrane (IC50 = 122 nM); this inhibition was observed in the absence of MgATP, and therefore was direct and unrelated to channel phosphorylation/dephosph orylation. 4. Channels in patches excised in outside-out mode were blo cked by 1 mar Ba2+ but were unaffected by 4-AP or TEA. 5. Channels in cell-attached patches were inhibited after single spikes, yielding inw ard ensemble currents lasting several hundred milliseconds. This was p revented in Ca2+-free solution, implying that it was due to Ca2+ entry . 6. The properties of these channels (block by internal Ca2+ and exte rnal Oxo-M and Ba2+ and the presence of both voltage-dependent and vol tage-independent components in their P-o/V relationship) show points o f resemblance to those expected for channels associated with, both I(l ea)k and I-K(M) components of the sustained macroscopic currents. For this reason we designate them K-sust ('sustained current') channels. I nhibition of these channels by Ca2+ entry during an action potential m ay account for some forms of Ca2+-dependent after depolarization. Thei r high sensitivity to internal Ca2+ may provide a new, positive feedba ck mechanism for cell excitation operating at low (near-resting) [Ca2](i).