Functional expression of Kir 6.1/SUR1-KATP channels in frog retinal Muller glial cells

The retinae and brains of larval and adult amphibians survive longlasting anoxia; this finding suggests the presence of functional K-ATP channels. We have previously shown with immunocytochemistry studies that retinal glial (Muller) cells in adult frogs express the K-ATP channel and receptor proteins, Kir6.1 and SUR1, while retinal neurons display Kir6.2 and SUR2A/B (Skatchkov et al., 2001a: NeuroReport 12:1437-1441; Eaton et al., in press: NeuroReport). Using both immunocytochemistry and electrophysiology, we demonstrate the expression of Kir6.1/SUR1 (K-ATP) channels in adult frog and tadpole Muller cells. Using conditions favoring the activation of K-ATP channels (i.e., ATP- and spermine-free cytoplasm-dialyzing solution containing gluconate) in Muller cells isolated from both adult frogs and tadpoles, we demonstrate the following. First, using the patch-clamp technique in whole-cell recordings, tolbutamide, a blocker of K-ATP, channels, blocks nearly 100% of the transient and about 30% of the steady-state inward currents and depolarizes the cell membrane by 5-12 mV. Second, inside-out membrane patches display a single-channel inward current induced by gluconate (40 mM) and blocked by ATP (200 muM) at the cytoplasmic side. The channels apparently show two sublevels (each of similar to27-32 pS) with a total of 85-pS maximal conductance at -80 mV; the open probability follows a two-exponential mechanism. Thus, functional K-ATP channels, composed of Kir6.1/SUR1, are present in frog Muller cells and contribute a significant part to the whole-cell K+ inward currents in the absence of ATP. Other inwardly rectifying channels, such as Kir4.1 or Kir2.1, may mediate the remaining currents. K-ATP, channels may help maintain glial cell functions during ATP deficiency. (C) 2002 Wiley-Liss, Inc.