Loss of Hyperpolarization-activated Cl− Current in Salivary Acinar Cells from Clcn2 Knockout Mice

Abstract

ClC-2 is localized to the apical membranes of secretory epithelia where it has been hypothesized to play a role in fluid secretion. Although ClC-2 is clearly the inwardly rectifying anion channel in several tissues, the molecular identity of the hyperpolarization-activated Cl ؊ current in other organs, including the salivary gland, is currently unknown. To determine the nature of the hyperpolarization-activated Cl ؊ current and to examine the role of ClC-2 in salivary gland function, a mouse line containing a targeted disruption of the Clcn2 gene was generated. The resulting homozygous Clcn2 ؊/؊ mice lacked detectable hyperpolarization-activated chloride currents in parotid acinar cells and, as described previously, displayed postnatal degeneration of the retina and testis. The magnitude and biophysical characteristics of the volume-and calcium-activated chloride currents in these cells were unaffected by the absence of ClC-2. Although ClC-2 appears to contribute to fluid secretion in some cell types, both the initial and sustained salivary flow rates were normal in Clcn2 ؊/؊ mice following in vivo stimulation with pilocarpine, a cholinergic agonist. In addition, the electrolytes and protein contents of the mature secretions were normal. Because ClC-2 has been postulated to contribute to cell volume control, we also examined regulatory volume decrease following cell swelling. However, parotid acinar cells from Clcn2 ؊/؊ mice recovered volume with similar efficiency to wild-type littermates. These data demonstrate that ClC-2 is the hyperpolarization-activated Cl ؊ channel in salivary acinar cells but is not essential for maximum chloride flux during stimulated secretion of saliva or acinar cell volume regulation.

A fluid secretion pathway unmasked by acinar-specific Tmem16A gene ablation in the adult mouse salivary gland

“…Ex vivo saliva secretion rates were measured in response to IPR in SMGs lacking ClC-2 channel (Clcn2 −/− ) and in WT glands treated with DCPIB and NPPB, two blockers of VRAC. IPR-induced saliva secretion was not affected in Clcn2 −/− glands, supporting previous reports that ClC-2 channels do not play a major role in saliva secretion (22,35). In contrast, IPR-induced saliva secretion was severely decreased by both DCPIB and NPPB.…”

“…S1). Gene targeting protocols for ACID, mT/mG (Jackson Laboratory), Clcn2 −/− , and Cftr ΔF508/ΔF508 mice were as described previously (11,14,22,40). Mice were housed in microisolator cages with ad libitum access to laboratory chow and water during 12-h light/dark cycles.…”

Leukoencephalopathy upon Disruption of the Chloride Channel ClC-2

“…It was most abundant in brain and in epithelia Cid et al, 1995). In mice, the disruption of ClC-2 led to testicular and retinal degeneration Nehrke et al, 2002). Germ cells and photoreceptors depend on the transepithelial transport mediated by Sertoli cells and the retinal pigment epithelium (RPE), respectively.…”

Evidence for a Functional Interaction between the ClC-2 Chloride Channel and the Retrograde Motor Dynein Complex

“…Recent studies in which ClC-2 expression was disrupted in model organisms support the claim that ClC-2 directly mediates this chloride conductance. For example, a chloride conductance path with the above biophysical properties was detected in studies of salivary gland cells obtained from normal mice whereas this function was absent in salivary gland cells obtained from ClC-2 knock-out mice (4). Similarly, depletion of the Caenorhabditis elegans ortholog of ClC-2, CLH-3, in maturing oocytes by RNA interference leads to the complete abrogation of the native, inwardly rectifying chloride conductance (5).…”