Chemicals inducing acute irritant contact dermatitis mobilize intracellular calcium in human keratinocytes.

Intracellular Ca(2+) increase is a common feature of multiple cellular pathways associated with receptor and channel activation, mediator secretion and gene regulation. We investigated the possibility of using this Ca(2+) signal as a biomarker for a reaction to chemical irritants of normal human keratinocytes (NHK) in submerged primary cell culture. We tested 14 referenced chemical compounds classified as strong (seven), weak (four) or non- (three) irritants in acute irritant contact dermatitis. We found that the strong irritant compounds tested at 20-40 mM induced an intracellular Ca(2+) increase measurable by spectrofluorimetry in an automated test. Weak and non-irritant compounds however did not increase intracellular Ca(2+) concentration. We further investigated the mechanisms by which the amine heptylamine, classified as a R34 corrosive compound, increases intracellular Ca(2+). Heptylamine (20mM) induced an ATP release that persisted in the absence of intra- and extra-cellular Ca(2+). In addition, we found that this ATP activates NHK purinergic receptors that subsequently cause the increase in intracellular Ca(2+) from sarcoplasmic reticular stores. We conclude that measuring the intracellular Ca(2+) concentration in NHK is a suitable and easy way of determining any potential reaction to soluble chemical compounds.

ATP signalling is crucial for the response of human keratinocytes to mechanical stimulation by hypo-osmotic shock.

Touch is detected through receptors located in the skin and the activation of channels in sensory nerve fibres. Epidermal keratinocytes themselves, however, may sense mechanical stimulus and contribute to skin sensation. Here, we showed that the mechanical stimulation of human keratinocytes by hypo-osmotic shock releases adenosine triphosphate (ATP) and increases intracellular calcium. We demonstrated that the release of ATP was found to be calcium independent because emptying the intracellular calcium stores did not cause ATP release; ATP release was still observed in the absence of external calcium and it persisted on chelating cytosolic calcium. On the other hand, the released ATP activated purinergic receptors and mobilized intracellular calcium stores. The resulting depletion of stored calcium led to the activation of capacitative calcium entry. Increase in cytosolic calcium concentration was blocked by the purinergic receptor blocker suramin, phospholipase C inhibitor and apyrase, which hydrolyses ATP. Collectively, our data demonstrate that human keratinocytes are mechanically activated by hypo-osmotic shock, leading first to the release of ATP, which in turn stimulates purinergic receptors, resulting in the mobilization of intracellular calcium and capacitative calcium entry. These results emphasize the crucial role of ATP signalling in the transduction of mechanical stimuli in human keratinocytes.

Mechanosensor Channels in Mammalian Somatosensory Neurons.

Mechanoreceptive sensory neurons innervating the skin, skeletal muscles andviscera signal both innocuous and noxious information necessary for proprioception, touchand pain. These neurons are responsible for the transduction of mechanical stimuli intoaction potentials that propagate to the central nervous system. The ability of these cells todetect mechanical stimuli impinging on them relies on the presence of mechanosensitivechannels that transduce the external mechanical forces into electrical and chemical signals.Although a great deal of information regarding the molecular and biophysical properties ofmechanosensitive channels in prokaryotes has been accumulated over the past two decades,less is known about the mechanosensitive channels necessary for proprioception and thesenses of touch and pain. This review summarizes the most pertinent data onmechanosensitive channels of mammalian somatosensory neurons, focusing on theirproperties, pharmacology and putative identity.