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Clinical Science (2007) 112, (167–174) (Printed in Great Britain)

Altered calcium signalling in platelets from bile-duct-ligated rats

Noemí M. ATUCHA*, David IYÚ*, Antonia ALCARAZ*, Vladimir ROSA*, Concepción MARTÍNEZ-PRIETO*, M. Clara ORTIZ*, Juan Antonio ROSADO† and Joaquín GARCÍA-ESTAÑ*

*Department of Physiology, Faculty of Medicine, University of Murcia, 30100 Murcia, Spain, and †Department of Physiology, University of Extremadura, Cáceres 10071, Spain

Key words: calcium signalling, bile-duct ligation, capacitative Ca²⁺ entry, cholestasis, liver cirrhosis, sarcoplasmic/endoplasmic-reticulum Ca²⁺-ATPase (SERCA), thrombin.

Abbreviations: 2-APB, 2-aminoethoxydiphenyl borate; AUC, area under the curve; BDL, bile-duct ligated; BDL group, rats at 4 weeks after bile-duct ligation; CCE, capacitative Ca²⁺ entry; cholestasis group, rats at 11–13 days after bile-duct ligation; fura-2/AM, fura 2 acetoxymethyl ester; IP₃, inositol 1,4,5-trisphosphate; PMCA, plasma membrane Ca²⁺-ATPase; SERCA, sarcoplasmic/endoplasmic-reticulum Ca²⁺-ATPase.

Correspondence: Professor Noemí M. Atucha (email ntma@um.es).

In the present study, we have analysed the mechanisms of Ca²⁺ entry and release in platelets obtained from BDL (bile-duct-ligated) rats, 11–13 days and 4 weeks after surgery. Platelets were washed and loaded with fura-2, and [Ca²⁺]_i (cytosolic Ca²⁺ concentration) was determined in cell suspensions by means of fluorescence spectroscopy. Basal [Ca²⁺]_i was similar in platelets from BDL rats compared with those from their respective controls, both in the absence and presence of extracellular Ca²⁺. Platelet stimulation with thrombin in the absence and presence of extracellular Ca²⁺ induced a rapid rise in [Ca²⁺]_i that was of greater magnitude in platelets from BDL rats than in controls. Ca²⁺ storage was significantly elevated in platelets from BDL rats, as well as the activity of SERCA (sarcoplasmic/endoplasmic-reticulum Ca²⁺-ATPase). Capacitative Ca²⁺ entry, as evaluated by inhibition of SERCA with thapsigargin, was also altered in platelets from BDL rats, having lower rates of Ca²⁺ entry. In conclusion, chronic BDL alters intracellular Ca²⁺ homoeostasis in platelets, such that an enhanced Ca²⁺ release is evoked by thrombin, which may be due to an increased amount of Ca²⁺ stored in the intracellular organelles and secondary to an enhanced activity of SERCA. These alterations are already evident before cirrhosis has completely developed and occurs during the cholestasis phase.

Received 18 August 2006; accepted 1 September 2006

Published as Immediate Publication 1 September 2006, doi:10.1042/CS20060226

©2007 The Biochemical Society