Nanoscale study of the as-grown hydrogenated amorphous silicon surface

A scanning tunneling microscope has been used to study the topography of the as-grown surface of device-quality, intrinsic, hydrogenated amorphous silicon deposited by rf discharge from silane. The substrates were atomically flat, oxide-free, single-crystal silicon or gallium arsenide. No evidence for island formation or nanoscale irregularities was seen in studies of 100-Å-thick films on either silicon or gallium arsenide. The topography of 1000- and 4000-Å-thick films has much variation; many regions can be characterized as rolling hills, but atomically flat areas have also been observed nearby. Generally, it appears that surface diffusion plays a role in smoothing the film topography. In most regions, the observed slopes were 10% or less from horizontal, but some steep-sided valleys, indicating incipient voids, were observed. The effect of the finite size of the scanning tunneling microscope probe tip is considered; this has an effect on the observed images in some cases

Prion Protein Misfolding Affects Calcium Homeostasis and Sensitizes Cells to Endoplasmic Reticulum Stress

Prion-related disorders (PrDs) are fatal neurodegenerative disorders characterized by progressive neuronal impairment as well as the accumulation of an abnormally folded and protease resistant form of the cellular prion protein, termed PrPRES. Altered endoplasmic reticulum (ER) homeostasis is associated with the occurrence of neurodegeneration in sporadic, infectious and familial forms of PrDs. The ER operates as a major intracellular calcium store, playing a crucial role in pathological events related to neuronal dysfunction and death. Here we investigated the possible impact of PrP misfolding on ER calcium homeostasis in infectious and familial models of PrDs. Neuro2A cells chronically infected with scrapie prions showed decreased ER-calcium content that correlated with a stronger upregulation of UPR-inducible chaperones, and a higher sensitivity to ER stress-induced cell death. Overexpression of the calcium pump SERCA stimulated calcium release and increased the neurotoxicity observed after exposure of cells to brain-derived infectious PrPRES. Furthermore, expression of PrP mutants that cause hereditary Creutzfeldt-Jakob disease or fatal familial insomnia led to accumulation of PrPRES and their partial retention at the ER, associated with a drastic decrease of ER calcium content and higher susceptibility to ER stress. Finally, similar results were observed when a transmembrane form of PrP was expressed, which is proposed as a neurotoxic intermediate. Our results suggest that alterations in calcium homeostasis and increased susceptibility to ER stress are common pathological features of both infectious and familial PrD models

ATP release via anion channels

ATP serves not only as an energy source for all cell types but as an ‘extracellular messenger-for autocrine and paracrine signalling. It is released from the cell via several different purinergic signal efflux pathways. ATP and its Mg2+ and/or H+ salts exist in anionic forms at physiological pH and may exit cells via some anion channel if the pore physically permits this. In this review we survey experimental data providing evidence for and against the release of ATP through anion channels. CFTR has long been considered a probable pathway for ATP release in airway epithelium and other types of cells expressing this protein, although non-CFTR ATP currents have also been observed. Volume-sensitive outwardly rectifying (VSOR) chloride channels are found in virtually all cell types and can physically accommodate or even permeate ATP4- in certain experimental conditions. However, pharmacological studies are controversial and argue against the actual involvement of the VSOR channel in significant release of ATP. A large-conductance anion channel whose open probability exhibits a bell-shaped voltage dependence is also ubiquitously expressed and represents a putative pathway for ATP release. This channel, called a maxi-anion channel, has a wide nanoscopic pore suitable for nucleotide transport and possesses an ATP-binding site in the middle of the pore lumen to facilitate the passage of the nucleotide. The maxi-anion channel conducts ATP and displays a pharmacological profile similar to that of ATP release in response to osmotic, ischemic, hypoxic and salt stresses. The relation of some other channels and transporters to the regulated release of ATP is also discussed

Modulation of intracellular pH by secretagogues and the Na+/H+ antiporter in cultured bovine chromaffin cells

The possible physiological role of cytosolic pH changes in adrenal medullary chromaffin cell secretion was examined by investigating the effects of catecholamine secretagogues on cytosolic pH, which was monitored using the intracellular fluorescent indicator 2′,7′-bis-(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF). Anti-fluorescein antibodies were used to reduce background fluorescence from extracellular 2′,7′-bis-(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF). Stimulation with both cholinergic agonists (acetylcholine, nicotine) and a depolarizing agent (high K+) transiently acidified the cytosol of the chromaffin cell. This acidification was antagonized by reducing extracellular Ca2+ concentration and by Ca2+ antagonists (Co2+, verapamil), indicating that it occurred secondarily to Ca2+ influx, possibly as a result of exchange of Ca2+ ions for protons across organelle membranes. Taken together with previously published data [Kuijpers G.A.J. et al. (1989) J. biol. Chem. 264, 698-7