Marie-Josée Duran

Specific Up-Regulation of Mitochondrial F0F1-ATPase Activity after Short Episodes of Atrial Fibrillation in Sheep

ATPase Activity and Atrial Fibrillation. Introduction: Ventricular fibrillation induced by either digitalis intoxication or electrical stimulation is reported to alter myocardial energy by impairing the sarcolemmal Na,K‐ATPase or the receptor for digitalis and the mitochondrial ATPase synthase or F0,F1‐ATPase. However, little is known about these membrane functions during atrial fibrillation (AF).

A Quantitative Immunocytochemical Study of Na+, K+-ATPase in Rat Hepatocytes After STZ-induced Diabetes and Dietary Fish Oil Supplementation

Because diabetes causes alterations in hepatic membrane fatty acid content, these changes may affect the Na+, K+-ATPase. In this study we documented the effects of streptozotocin (STZ)-induced diabetes on hepatic Na+, K+-ATPase catalytic α1-subunit and evaluated whether these changes could be normalized by fish oil supplementation. Two groups of diabetic rats received fish oil or olive oil supplementation. Both groups had a respective control group. We studied the localization of catalytic α1-subunit on bile canalicular and basolateral membranes using immunocytochemical methods and confocal laser scanning microscopy, and the Na+, K+-ATPase activity, membrane fluidity, and fatty acid composition on isolated hepatic membranes. A decrease in the α1-subunit was observed with diabetes in the bile canalicular membranes, without changes in basolateral membranes. This decrease was partially prevented by dietary fish oil. Diabetes induces significant changes as documented by enzymatic Na+, K+-ATPase activity, membrane fluidity, and fatty acid content, whereas little change in these parameters was observed after a fish oil diet. In conclusion, STZ-induced diabetes appears to modify bile canalicular membrane integrity and dietary fish oil partly prevents the diabetes-induced alterations.

Critical Role of the Isoform-Specific Region in α1-Na,K-ATPase Trafficking and Protein Kinase C-Dependent Regulation

The isoform-specific region (ISR) is a region of structural heterogeneity among the four isoforms of the catalytic alpha-subunit of the Na,K-ATPase and an important structural determinant for isoform-specific functions. In the present study, we examined the role of a potential dileucine clathrin adaptor recognition motif [DE]XXXL[LI] embedded within the alpha1-ISR. To this end, a rat alpha1 construct where leucine 499 was replaced by a valine (as found in the alpha2 isoform sequence) was compared to wild-type rat alpha1 after stable expression in opossum kidney cells. Total Na,K-ATPase expression, activity, or in situ (86)Rb(+) transport was not affected by the L499V mutation. However, surface Na,K-ATPase expression was nearly doubled. This increase was associated with a reduced rate of internalization from the cell surface of about 50% after a 4 h chase and became undetectable if clathrin-coated pit-mediated trafficking was blocked with chlorpromazine. Further, PKC-induced stimulation of Na,K-ATPase-mediated (86)Rb(+) uptake was doubled in mutant-expressing cells, comparable to the chimera containing the intact alpha2-ISR. Similar results were observed when the potential motif was disrupted by means of an E495S mutation. These findings suggest that a dileucine motif embedded within the Na,K-ATPase alpha1-ISR plays a critical role in the surface expression of Na,K-ATPase alpha1 polypeptides at steady state and in the response to PKC activation.

REMODELING OF Na,K-ATPase, AND MEMBRANE FLUIDITY AFTER ATRIAL FIBRILLATION IN SHEEP

ABSTRACT Atrial fibrillation (AF) is accompanied by various changes in ion channels that cause atrial electrophysiological remodeling. The enzyme Na,K-ATPase is also a major cellular mechanism for the regulation of ion homeostasis. During AF, Na,K-ATPase may be regulated by synthesis of its α- and β-subunits as well as changes in membrane fluidity. To test this hypothesis, we studied the effect of pacing-induced AF in sheep on atrial Na,K-ATPase α- and β-subunits and on membrane fluidity as well. Methods: A group of six sheep (AF group) was subjected to overdrive electrical stimulation of the right atrium in order to induce AF. A group of six sham operated sheep served as control. All paced sheep developed multiple episodes of sustained AF with a mean total duration of 110 min over a 2-hours period. Protein expression of Na,K-ATPase α- and β-subunits in atrial microsomal membranes was assayed by Western blotting analysis. When significant changes in membrane expression were observed, transcriptional regulation was analysed by Northern blotting. Membrane fluidity was assessed on atrial microsomal fractions by anisotropy measurements using the fluorescent probe diphenylhexatriene. Results: Atrial fibrillation enhanced the expression of the Na,K-ATPase β1-subunit at both membrane and mRNA levels. Anisotropy values were higher in AF group than in control group, indicating a decreased fluidity of the membranes isolated from paced sheep atria. Conclusion: These data are the first evidence for an enhanced Na,K-ATPase β1-subunit expression in membrane during AF. Membrane rigification represents a new factor of tachycardia-induced atrial remodeling.