Biomedical subjects

A Bryan-Sisneros

Publications and source records attributed to A Bryan-Sisneros.

3 recordsLinked to original sources

PubMed2000

Dual role of ATP in supporting volume-regulated chloride channels in mouse fibroblasts.

The effects of inhibitors of protein tyrosine kinases (PTKs) on the Cl(-) current (I(Cl(vol))) through volume-regulated anion/chloride (VRAC) channels whilst manipulating cellular ATP have been studied in mouse fibroblasts using the whole-cell patch clamp technique. Removal of ATP from the pipette-filling solution prevented activation of the current during osmotic cell swelling and when the volume of patched cells was increased by the application of positive pressure through the patch pipette to achieve rates exceeding 100%/min. Equimolar substitution of ATP in the pipette solution with its non-hydrolyzable analogs, adenosine 5'-O-(3-thiotriphosphate) (ATPgammaS) or adenylyl-(beta,gamma-methylene)-diphosphonate (AMP-PCP), not only supported activation of the current but also maintained its amplitude. The PTK inhibitors, tyrphostins A25, B46, 3-amino-2,4-dicyano-5-(4-hydroxyphenyl)penta-2,4-dienonitrile++ + and genistein (all at 100 microM), inhibited I(Cl(vol)) in a time-dependent manner. Tyrphostin A1, which does not inhibit PTK activity, did not affect the current amplitude. The PTK inhibitors also inhibited I(Cl(vol)) under conditions where ATP in the pipette was substituted with ATPgammaS or AMP-PCP. We conclude that in mouse fibroblasts ATP has a dual role in the regulation of the current: it is required for protein phosphorylation to keep VRAC channels operational and, through non-hydrolytic binding, determines the magnitude of I(Cl(vol)). We also suggest that tyrosine-specific protein kinases and phosphatases exhibit an interdependent involvement in the regulation of VRAC channels.

Adenosine Triphosphate↗

PubMed1999

A functional spliced-variant of beta 2 subunit of Kv1 channels in C6 glioma cells and reactive astrocytes from rat lesioned cerebellum.

Voltage-gated K(+) channels (Kv1) are important in glia, being required for cell proliferation. Herein, reactive astrocytes from a rat cerebellar lesion were shown to contain Kv1.1, -1.2, -1.3, -1.4, and -1.6 alpha plus beta1.1 subunits, as well as an unusual beta2.1 constituent; the latter was also found in a glioblastoma C6 cell line, together with Kv1.1, -1.3, and -1.6 and beta1.1 subunits. Reverse transcriptase-polymerase chain reaction revealed a novel product of the beta2 gene in C6 cells and reactive astrocytes, but not in cultured astrocytes or rat normal brain. Its cloning identified a variant, Kvbeta2.1A, alternatively spliced between I24 and Y39. Despite this 14 residue deletion, Kvbeta2.1A assembled cotranslationally with Kv1.1 or -1.2 and, when coexpressed with Kv1. 4 in oocytes, increased the inactivation rate of this K(+) current. Whereas the full-length beta2.1 gave a large increase in the amplitude of the Kv1.1 current in oocytes, the effect of beta2.1A varied from a modest elevation of the current to a slight suppression in some cases. In summary, this is the first report of the existence of an alternatively spliced product of the Kvbeta2.1 gene in C6 cells and reactive astrocytes, and supports the involvement of its core region (residues 39 onward) in assembly with alpha subunits while excluding a contribution of the adjacent 14 residues to accelerating the inactivation of Kv1.4.

Alternative Splicing↗

PubMed1999

Protein phosphotyrosine phosphatase inhibitors suppress regulatory volume decrease and the volume-sensitive Cl- conductance in mouse fibroblasts.

The effects of the protein tyrosine phosphatase (PTP) inhibitors, pervanadate, monoperoxo(picolinato)- oxo-vanadate(V) [mpV(pic)] and dephostatin, on regulatory volume decrease (RVD) and the volume-sensitive Cl- current in mouse L-fibroblasts were studied with the aid of video microscopy and the whole-cell patch-clamp technique. The RVD induced by the hyposmotic shift from 300 to 150 mosmol/l, was strongly suppressed in cells that had been pre-incubated in pervanadate (25 microM) or in mpV(pic) (10 microM), or subjected to extracellular application of dephostatin (20 microM). The acceleration in RVD caused by gramicidin (0.5 microM) was also slowed down by pervanadate pre-treatment, suggesting that the PTP inhibitors affected the volume-sensitive Cl- conductance. Inhibition of the volume-sensitive Cl- current by pervanadate (25 microM) pre-treatment and by acutely applied dephostatin (20 microM) was confirmed in the whole-cell experiments (by @70% and by @50%, respectively). Both pervanadate and dephostatin inhibited the outward and inward Cl- currents equally, which suggests that only the number of open channels was affected. The amplitude of the Cl- current decreased slowly during application of dephostatin and did not recover after its termination. We conclude that in mouse L-fibroblasts, similar to bovine chromaffin cells, inhibition of PTPs results in the suppression of both RVD and the volume-sensitive Cl- current.

Animals↗