Electron probe X-ray microanalysis of cellular ions in the eccrine secretory coil cells during methacholine stimulation

Abstract

Intracellular concentrations of Na, K, Cl ([Na], [K] and [Cl], respectively) and other elements were determined in isolated monkey eccrine sweat secretory coil cells using quantitative electron probe X-ray microanalysis of freeze dried cryosections. The validity of the methodology was partially supported by qualitative agreement of the X-ray microanalysis data with those obtained by micro-titration with a helium glow spectrophotometer. [Na], [K] and [Cl] of the cytoplasm were the same as those in the nucleus in both clear and dark cells. [Na], [K], and [Cl] of the clear cells were also the same as those of the dark cells at rest and after stimulation with methacholine (MCh), suggesting that these two cell types behave like a functional syncytium. MCh stimulation induced a pharmacologically specific, dose-dependent decrease in [K] and [Cl] (as much as 65%), and a 3.7-fold increase in [Na]. In myoepithelial cells, a similar change in [Na] and [K] was noted after MCh stimulation although the decrease in [Cl] was only 20%. The MCh-induced change in [Na], [K] and [Cl] was almost completely inhibited by removal of Ca2+ from the medium. 10(-4) M bumetanide inhibited the MCh-induced increase in [Na], reduced the decrease in [K] by about 50%, but slightly augmented the MCh-induced decrease in [Cl]. 10(-4) M ouabain increased [Na] and decreased [K] as did MCh; however, unlike MCh, ouabain increased [Cl] by 56% after 30 min of incubation. Thus the data may be best interpreted to indicate that Ca-dependent K efflux and (perhaps also Ca-dependent) Cl efflux are the predominant initial ionic movement in muscarinic cholinergic stimulation of the eccrine sweat secretory coils and that the ouabain-sensitive Na pump plays an important role in maintenance of intracellular ions and sweat secretion.

Application of Cationic Colloidal Gold Staining for the Demonstration of Anionic Sites.

“…These results are consistent with the previous report of xray microanalysis that has shown the presence of a high concentration of calcium in the dark cell granules [26].…”

Quantitative ion determination in eccrine sweat gland cells correlates to sweat reduction of antiperspirant actives

“…Interestingly, our observed influences of the ion equilibrium evoked by sweat‐reducing agents are the exact opposite of the reactions described by Saga & Sato in sweat‐stimulated native eccrine ape sweat glands. In their study, X‐ray microanalysis of isolated, methacholine stimulated ape sweat gland coil cells detected an increase in [Na + ] i while [Cl − ] i was reduced [43]. These opposing impacts of the stimulator methacholine, on the one hand, and the modulation by the inhibitors ACH and PEI‐HCl, on the other hand, on sweat‐relevant ions corroborate the hypothesis of biochemically steered sweating.…”

A novel cell line from human eccrine sweat gland duct cells for investigating sweating physiology

“…As our primary intention of establishing an immortalized eSG duct-derived cell line was to apply it in in vitro screening assays for identification of alternative antiperspirant actives, their reactivity towards substance application was also tested. To achieve this, the approach of loading ion-sensitive fluorescence dyes into the cells was preferred over earlier in research used patch-clamp technique [72] and X-ray microanalysis [51,[73][74][75] as the latter methods are much more complex and require more sophisticated equipment.…”