Volume regulatory decrease mediated by extracellular nucleotides.

PAFUNDO, D. E., CHARA, O. Y SCHWARZBAUM, P. J.

Montevideo, Uruguay.

Simposio; VI conferencia internacional de física biológica, V congreso de biofísica del cono sur y XXXIV reunión anual de la Sociedad de Biofísica Argentina; 2007

Nearly all cell types possess mechanisms that enable the release of ATP in response to an hypotonic challenge. Following cell swelling, extracellular ATP (ATPe) activate specific P receptors that allow the cell to downregulate its volume towards isotonic values. This work aimed at studying the factors governing the kinetics of ATPe of hypotonically exposed goldfish hepatocytes. Under hypotonicity the concentration of ATPe is mainly governed by lytic and non-lytic release of the nucleotide, hydrolysis of ATP by ecto-ATPase activity at the cell surface and ATP diffusion within the extracellular compartment. We found that goldfish hepatocytes release ATP after hypotonic shock. The time course of ATPe is non-monotonic showing a maximum at 725±165 nM (106 cells)-1. In order to simulate the kinetics of ATPe we developed a one dimensional mathematical model with three compartments: the intracellular (i), an extracellular near to the cells membrane (e1), and one representing the bulk extracellular medium (e2). ATPe is controlled by: 1) non-lytic release of ATP (JNL), 2) lytic ATP release, 3) Ecto‑ATPase activity in e1, 4) ATP diffusion between e1 and e2. JNL was described as a function JR multiplied by a function F accounting for a positive feedback mechanism. Four different JR were tested; i) constant JR, ii) step function (JR is zero until it becomes activated and remains constant thereafter); iii) impulse function (JR has the form of a rectangular pulse that can be triggered and shut off at variable times); iV) a log-normal function which includes a non-linear fast increase to a maximum, followed by a relatively slowly non-linear decrease. Use of JR functions i-iv did not allow the model to provide a good fit to experimental data. However, using the log-normal function we simulated the kinetics of ATPe with reasonable accuracy. The predicted JNL showed a 1.93 10-18 mol sec-1 peak after 2 sec. The model was also used to quantify the relative importance of processes 1-4 in determining the kinetics of ATPe.