Biomedical subjects

Andrew Shevchuk

Publications and source records attributed to Andrew Shevchuk.

4 recordsLinked to original sources

PubMed2005

Aldosterone acts via an ATP autocrine/paracrine system: the Edelman ATP hypothesis revisited.

Aldosterone, the most important sodium-retaining hormone, was first characterized >50 years ago. However, despite numerous studies including the classical work of Isidore S. "Izzy" Edelman showing that aldosterone action depended on ATP production, the mechanism by which it activates sodium reabsorption via the epithelial sodium channel remains unclear. Here, we report experiments that suggest that one of the key steps in aldosterone action is via an autocrine/paracrine system. The hormone stimulates ATP release from the basolateral side of the target kidney cell. Prevention of ATP accumulation or its removal blocks aldosterone action. ATP then acts via a purinergic mechanism to produce contraction of small groups of adjacent epithelial cells. Patch clamping demonstrates that it is these contracted cells that have channel activity. With progressive recruitment of contracting cells, there is then a parallel increase in transepithelial electrical conductance. In common with other stimuli of sodium transport, this pathway involves phosphatidylinositol 3-kinase. Inhibition of phosphatidylinositol 3-kinase blocks both cell contraction and conductance. We put forward the hypothesis that redistribution of the cell volume caused by the lateral contraction results in apical swelling and that this change, in turn, disrupts the epithelial sodium channel interaction with the F-actin cytoskeleton, opening the channel and hence increasing sodium transport.

Adenosine Triphosphate↗

PubMed2005

The scanned nanopipette: a new tool for high resolution bioimaging and controlled deposition of biomolecules.

The boundary between the physical and biological sciences has been eroded in recent years with new physical methods applied to biology and biological molecules being used for new physical purposes. We have pioneered the application of a form of scanning probe microscopy based on a scanned nanopipette, originally developed by Hansma and co-workers, for reliable non-contact imaging over the surface of a live cell. We have found that the nanopipette can also be used for controlled local voltage-driven application of reagents or biomolecules and this can be used for controlled deposition and the local delivery of probes for mapping of specific species. In this article we review this progress, focussing on the physical principles and new phenomena that we have observed, and then outline the future applications that are now possible.

Aniline Compounds↗

PubMed2005

Scanning ion conductance microscopy reveals how a functional renal epithelial monolayer maintains its integrity.

BACKGROUND: To function as a transport barrier a renal tubule epithelial monolayer needs to maintain its integrity when, sudden hypertonic stress causes cell shrinkage, new cells are added, or cells in the monolayer die. However, the mechanism used to achieve this is largely unknown. Scanning ion conductance microscopy (SICM) has been shown to be suitable for imaging the surface of live renal cells with high topographic resolution, and can be used to elucidate how a functional renal epithelial monolayer maintains its integrity. METHODS: SICM was used for high spatial resolution topographic imaging of Xenopuslaevis renal epithelial A6 cells cultured on membrane filter inserts. RESULTS: The SICM images of A6 cells showed that the epithelial monolayer maintains its integrity under hypertonic stress, and during cell division and death. Sequential SICM topographic images revealed detailed structural changes and their time course for these protective processes, which involve highly cooperative cell movement. Some "balloon-like" structures were observed at susceptible tight junction regions, which were proposed to help cell maintaining the monolayer permeability integrity. CONCLUSION: SICM is a powerful tool for research on living renal epithelial cells, and has been used to elucidate how a functional epithelial monolayer maintains its integrity. Using this technique we have observed that during hypertonic stress and regeneration, an organized sequence of events protect the loss of integrity of monolayer so that tight junctions and cell-cell contact are maintained and disruption to the function of whole monolayer is prevented.

Animals↗

PubMed2002

High-resolution scanning patch-clamp: new insights into cell function.

Cell specialization is often governed by the spatial distribution of ion channels and receptors on the cell surface. So far, little is known about functional ion channel localization. This is due to a lack of satisfactory methods for investigating ion channels in an intact cell and simultaneously determining the channels' positions accurately. We have developed a novel high-resolution scanning patch-clamp technique that enables the study of ion channels, not only in small cells, such as sperm, but in submicrometer cellular structures, such as epithelial microvilli, fine neuronal dendrites, and, particularly, T-tubule openings of cardiac myocytes. In cardiac myocytes, as in most excitable cells, action potential propagation depends essentially on the properties of ion channels that are functionally and spatially coupled. We found that the L-type calcium and chloride channels are distributed and colocalized in the region of T-tubule openings, but not in other regions of the myocyte. In addition, chloride channels were found in narrowly defined regions of Z-grooves. This finding suggests a new synergism between these types of channels that may be relevant for action potential propagation along the T-tubule system and excitation-contraction coupling.

Animals↗