This work aimed to study the relationship between agonist-induced changes in cytosolic free calcium levels, protein kinase C (PKC) activity and intracellular pH in isolated liver cells. We observed that, like alpha1-adrenergic agonists, the Ca2+-mobilizing vasoactive peptides vasopressin and angiotensin II produced an extracellular-Na+-dependent, 5-(N-ethyl-N-isopropyl)amiloride-sensitive, intracellular alkalinization, indicative of Na+/H+ antiporter activation. Blocking the agonist-induced increase in the intracellular Ca2+ concentration using the calcium chelator bis-(o-aminophenoxy)ethane-N,N,N', N'-tetra-acetic acid (BAPTA) prevented all types of receptor-mediated intracellular alkalinization. Thus activation of the Na+/H+ exchanger by either alpha1-adrenergic agonists or vasoactive peptides relies on the mobilization of intracellular Ca2+. In contrast, only the alpha1-adrenergic-agonist-induced alkalinization was dependent on extracellular Ca2+. Even though alpha1-adrenergic as well as vasoactive peptide agonists stimulated protein kinase C (PKC) activity in isolated liver cells, only the alpha1-adrenoreceptor-mediated intracellular alkalinization was dependent on PKC. According to these observations, Ca2+-mobilizing agonists appear to activate the Na+/H+ exchanger by at least two different mechanisms: (1) the alpha1-adrenoreceptor-mediated activation that is dependent on extracellular Ca2+ and PKC; and (2) vasoactive-peptide-induced alkalinization that is independent of extracellular Ca2+ and PKC. The alpha1-adrenoreceptor-mediated, PKC-sensitive, activation of the Na+/H+ exchanger seems to be responsible for the distinct ability of these receptors to elicit the sustained stimulation of hepatic functions.

The present work aimed to elucidate the influence of thyroid functional status on the α1-adrenoreceptor-induced activation of hepatic metabolic functions. The experiments were performed in either a nonrecirculating liver perfusion system featuring continuous monitoring of portal pressure,[Formula: see text], pCa, and pH, or isolated hepatocytes from euthyroid, hyperthyroid, and hypothyroid rats. Hypothyroidism decreased the α1-adrenergic stimulation of respiration, glycogen breakdown, and gluconeogenesis. These effects were accompanied by a decreased intracellular Ca2+ mobilization corroborating that those processes are regulated by the Ca2+-dependent branch of the α1-adrenoreceptor signaling pathway. Moreover, in hyperthyroid rats the α1-adrenergic-induced increase in cytosolic Ca2+ was enhanced, and glucose synthesis or mobilization was not altered. The thyroid status influenced neither the α1-adrenergic stimulation of vascular smooth muscle contraction nor the α1-agonist-induced intracellular alkalinization and protein kinase C (PKC) activation. Thus the distinct impairment of the Ca2+-dependent branch of the α1-adrenoreceptor signaling pathway by thyroid status provides a useful tool to investigate the role played by each signaling pathway, Ca2+ or PKC, in controlling hepatic functions.

This work aimed to investigate the acute effect of the thyroid hormone 3,5,3'-tri-iodo-L-thyronine (T3) in regulating the hepatic metabolism either directly or by controlling the responsiveness to Ca2+-mobilizing agonists. We did not detect any acute metabolic effect of T3 either in perfused liver or in isolated liver cells. However, T3 exerted a powerful inhibitory effect on the alpha1-adrenoreceptor-mediated responses. The promptness of this T3 effect rules out that it was the result of rate changes in gene(s) transcription. T3 inhibited the alpha1-adrenoreceptor-mediated sustained stimulation of respiration and release of Ca2+ and H+, but not the glycogenolytic or gluconeogenic responses, in perfused liver. In isolated liver cells, T3 enhanced the alpha1-agonist-induced increase in cytosolic free Ca2+ and impeded the intracellular alkalinization. Since T3 also prevented the alpha1-adrenoreceptor-mediated activation of protein kinase C, its effects on pH seem to be the result of a lack of activation of the Na+/H+ exchanger. The failure of T3 to prevent the alpha1-adrenergic stimulation of gluconeogenesis despite the inhibition of protein kinase C activation indicates that the elevation of cytosolic free Ca2+ is a sufficient signal to elicit that response. T3 also impaired some of the angiotensin-II-mediated responses, but did not alter the effects of PMA on hepatic metabolism, indicating, therefore, that some postreceptor event is the target for T3 actions. The differential effect of T3 in enhancing the alpha1-adrenoreceptor-mediated increase in cytosolic free Ca2+ and preventing the activation of protein kinase C, provides a unique tool for further investigating the role of each branch of the signalling pathway in controlling the hepatic functions. Moreover, the low effective concentrations of T3 (<= 10 nM) in perturbing the alpha1-adrenoreceptor-mediated response suggests its physiological significance.