Single‐channel characteristics of the large‐conductance anion channel in rat cortical astrocytes in primary culture

Abstract

Cultured rat cortical astrocytes, in addition to a variety of voltage-sensitive potassium channels, also express anion channels. However, the behavior and regulation of these anion channels have been far less studied. This paper describes a patch-clamp study on a voltage-sensitive 200-300 pS high-conductance single-channel anion current, which seems to possess at least five different open sublevels or, alternatively, be formed from five or more small-conductance ion channels linked together. This channel is voltage dependent, showing a bell-shaped open probability curve with highest open probability close to the reversal potential (zero-current). Although potassium channels are commonly detected in astrocytes in cell-attached and excised patches with both normal osmolarity and hypoosmotic solutions, the occurrence of the anion channel is clearly increased in isolated patches when hypoosmotic bath solutions are used. Also, cell aging in culture and the preparation of secondary cell cultures by trypsinization seem to increase the rate of occurrence of the anion channel. Though this channel is more routinely seen when a membrane patch is excised from the cell, occasionally cell-attached configurations with instant channel activity can be formed. While the modulation of this anion channel was being studied, it was found to be blocked by an anion transport inhibitor, L-644,711, reported to affect cell volume regulation in astrocytes.

The maxi-anion channel: a classical channel playing novel roles through an unidentified molecular entity

“…Modified from Sabirov et al [8] 2 day old rats [18] and adult humans [47] as well as in freshly dissected rat spinal root Schwann cells [48]. These channels were similar to those observed in cultured cortical astrocytes from rats [4,49,50] and mice [5][6][7]51] as well as in a rat astrocytic RGCN cell line [52]. In epithelia, the urinary bladder [53], gastric [54], pancreatic [55][56][57], colonic [58][59][60], airway [61][62][63], choroid plexus [64], bile duct [65,66], ciliary [67][68][69], renal [9,19,[70][71][72][73][74][75][76][77][78], vestibular [79], placental [80][81][82][83][84][85][86], ruminal …”

“…5), which blocks not only CFTR but also VSOR [122], in mouse C127 cells [8], rat cardiomyocytes [2] and mouse astrocytes [5]. An anion channel inhibitor, L-644-711 (0.5-1 mM), also blocked maxi-anion channels in cultured rat astrocytes [4]. Arachidonic acid at a micromolar level inhibited maxi-anion channels in an L6 rat muscle cell line [26], from human term placentas membranes reconstituted into giant liposomes [84], in mouse mammary C127 cells [112], in cultured neonatal rat cardiomyocytes [2,12] (Fig.…”

“…Consistent with the phenotype of the whole-cell VSOR current, this channel current exhibits outward rectification and voltage-dependent inactivation at large positive potentials ([?50 to ?100 mV). However, many authors have also reported the single-channel event with a much larger unitary conductance (300-400 pS) observed in the cell-attached mode after cell swelling [2][3][4][5][6][7][8][9]. A representative example of the maxianion channel currents recorded in the cell-attached patch on an osmotically swollen mammary C127 cells is shown in Fig.…”

Chloride/anion channels in glial cell membranes

“…This was seen as a clear sign of negligible chloride permeability; any subsequent experiments with these glial cells were conducted in low-chloride solutions. Also, investigators undertaking patch-clamp measurements with normal cultured astrocytes did not find any evidence of a significant chloride current activity at resting potentials (Bevan et al, 1985;Sonnhof, 1987;Barres et al, 1990;Jalonen, 1993;Lascola and Kraig, 1996). Consequently, this issue was ignored in many studies, and investigators simply assumed a negligible chloride conductance for all types of glial cells.…”

“…Another channel commonly found in cultured astrocytes is a large anion conductance, which has been known for quite some time (Bevan et al, 1985;Nowak et al, 1987;Sonnhof, 1987;Jalonen, 1993). It has a peculiar property of only being found in excised patches, as if an inhibitory factor prevents expression in intact cells.…”