title: Chronic hypocalcemia of vitamin D deficiency leads to lower intracellular calcium concentrations in rat hepatocytes.
abstract: Several lines of evidence indicate that calcium deficiency is associated with cellular defects in many tissues and organs. Owing to the large in vivo gradient between ionized extra- and intracellular Ca2+ concentrations ([Ca2+]i), it is generally recognized that the prevailing circulating Ca2+ does not significantly affect resting cytosolic Ca2+. To probe the consequences of hypocalcemia on [Ca2+]i, a model of chronic hypocalcemia secondary to vitamin D (D) deficiency was used. Hepatocytes were isolated from livers of hypocalcemic D-deficient, of normocalcemic D3-repleted, or of normal control rats presenting serum Ca2+ of 0.78 +/- 0.02, 1.24 +/- 0.03, or 1.25 +/- 0.01 mM, respectively (P < 0.0001). [Ca2+]i was measured in cell couplets using the fluorescent probe Fura-2. Hepatocytes of normocalcemic D3-repleted and of normal controls exhibited similar [Ca2+]i of 227 +/- 10 and 242 +/- 9 nM, respectively (NS), whereas those of hypocalcemic rats had significantly lower resting [Ca2+]i (172 +/- 10 nM; P < 0.0003). Stimulation of hepatocytes with the alpha 1-adrenoreceptor agonist phenylephrine illicited increases in cytosolic Ca2+ leading to similar [Ca2+]i and phosphorylase a (a Ca(2+)-dependent enzyme) activity in all groups but in contrast to normocalcemia, low extracellular Ca2+ was often accompanied by a rapid decay in the sustained phase of the [Ca2+]i response. When stimulated with the powerful hepatic mitogen epidermal growth factor (EGF), hepatocytes isolated from hypocalcemic rat livers responded with a blunted maximal [Ca2+]i of 237.6 +/- 18.7 compared with 605.2 +/- 89.9 nM (P < 0.0001) for their normal counterparts, while the EGF-mediated DNA synthesis response was reduced by 50% by the hypocalcemic condition (P < 0.03). Further studies on the possible mechanisms involved in the perturbed [Ca2+]i homeostasis associated with chronic hypocalcemia revealed the presence of an unchanged plasma membrane Ca2+ ATPase but of a significant decrease in agonist-stimulated Ca2+ entry as indicated using Mn2+ as surrogate ion (P < 0.03). Our data, thus indicate that, in rat hepatocytes, the in vivo calcium status significantly affects resting [Ca2+]i, and from this we raise the hypothesis that this lower than normal [Ca2+]i may be linked, in calcium disorders, to inappropriate cell responses mediated through the calcium signaling pathway as illustrated by the response to phenylephrine and EGF.
authors.affiliation: Centre de Recherche Clinique André-Viallet, Hôpital Saint-Luc, Montreal, Canada.
authors.forename: M
authors.initials: M
authors.lastname: Gascon-Barré
authors.forename: P
authors.initials: P
authors.lastname: Haddad
authors.forename: S J
authors.initials: SJ
authors.lastname: Provencher
authors.forename: S
authors.initials: S
authors.lastname: Bilodeau
authors.forename: F
authors.initials: F
authors.lastname: Pecker
authors.forename: S
authors.initials: S
authors.lastname: Lotersztajn
authors.forename: S
authors.initials: S
authors.lastname: Vallières
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D002762:Cholecalciferol / Q000494:pharmacology; D004815:Epidermal Growth Factor / Q000494:pharmacology; D006706:Homeostasis / Q000502:physiology; D006996:Hypocalcemia / Q000209:etiology / Q000378:metabolism*; D008099:Liver / Q000166:cytology / Q000378:metabolism*; D008297:Male; D010656:Phenylephrine / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D014808:Vitamin D Deficiency / Q000150:complications / Q000378:metabolism*
chemical_list: D002762:Cholecalciferol; D010656:Phenylephrine; D004815:Epidermal Growth Factor; D002118:Calcium