Characterization of monovalent ion transport systems in an insect cell line (Manduca sexta embryonic cell line che)

Abstract

The monovalent ion transport systems of an immortalized insect cell line (CHE) have been investigated. These cells are unusual in that unlike most vertebrate cells, their normal extracellular environment consists of high potassium and low sodium concentrations. CHE cells maintained high intracellular [K+] through both a furosemide-inhibitable and a vanadate-inhibitable transport system. Intracellular exchangeable [Na+] was slightly lower than the extracellular [Na+] and was maintained at this level through a vanadate-sensitive transport system. Na+ uptake was also inhibited by furosemide: however, the stoichiometry of furosemide-sensitive Na+ uptake when compared with furosemide-sensitive K+ uptake indicated that these cations are not cotransported. 4,4'-Diisothiocyano-2,2'-disulfonic acid stilbene (DIDS) inhibited Na+, K+, and Cl- uptake. Vanadate and furosemide decreased cytoplasmimic pH, while cytoplasmic pH increased in the presence of DIDS. A model is presented explaining how Na+, K+, Cl-, H+ and HCO3-fluxes are regulated in these cells.

Toxicity and disruption of midgut physiology in larvae of the European corn borer, Ostrinia nubilalis, by anion transporter blockers

“…A positive correlation between midgut alkalinity decrease and Cl À transport inhibition in this investigation suggested the observed alkalinity decrease was due to the inhibition of Cl À /HCO 3 -exchange between the midgut lumen and epithelia. In a previous study also, DIDS at 100 mM was shown blocking 36 Cl À exchange in M. sexta embryo cell culture (English and Cantley, 1984).…”

Characterization of chloride uptake in Drosophila Kc cells

“…A plasma membrane ATPdependent C1-pump has been described in such diverse cells as human neutrophils and epithelium from the locust hindgut (Simchowitz and de Weer, 1986;Hanrahan and Phillips, 1982). In Manduca sexta CHE cells, Cl-/HC03-exchange has been shown to be indirectly linked to the functioning of a vanadate-sensitive K+/Hf ATPase (English and Cantley, 1984). Under the conditions we have tested, the [Cl-li is as much as fourfold higher than the [Cl-I,.…”

Delta endotoxin inhibits Rb+ uptake, lowers cytoplasmic pH and inhibits a K+-ATPase inManduca sexta CHE cells

“…We prevb ously demonstrated that Rb + adequately substituted for K + in these insect cells (English & Cantley, 1984). Uptake assays were performed according to the method described by Smith et a[.…”

“…CHE cells have vanadate-inhibitable 86Rb+ uptake that is coincident with a vanadate-inhibitable cellular pH regulatory system, possibly resulting from coupled K+,H+-ATPase (English & Cantley, 1984). The cells maintain a small Na* gradient possibly due to a ouabain-sensitive ATPase implied by ouabain-sensitive ,8-[2-furyl] acryloyl phosphatase activity in plasma membranes (English & Marks, 1981).…”

“…The cells maintain a small Na* gradient possibly due to a ouabain-sensitive ATPase implied by ouabain-sensitive ,8-[2-furyl] acryloyl phosphatase activity in plasma membranes (English & Marks, 1981). Also, CHE cells have two distinct HCO3-regulatory systems: a CI-/HCO~-exchanger sensitive to 4,4'-diisothiocyano-2,2'-disulfonic acid stilbene (DIDS) and a furosemide and D1DS-sensitive KC[/ H +, HCO2 exchange system (English & Cantley, 1984). This communication relates that DET inhibits the vanadate-sensitive 8~Rb+ uptake and induces a reduction in cytoplasmic pH in whole CHE cells.…”