Roles of Ca²⁺ and the Ca²⁺‐sensing receptor (CASR) in the expression of inducible NOS (nitric oxide synthase)‐2 and its BH₄ (tetrahydrobiopterin)‐dependent activation in cytokine‐stimulated adult human astrocytes

Abstract

Since NO production by NOS-2 made by astrocytes activated by proinflammatory cytokines contributes to the killing of neurons in variously damaged human brains, knowing the mechanisms responsible for NOS-2 expression should contribute to developing effective therapeutics. The expression and activation of NOS-2 in normal adult human cerebral cortical astrocytes treated with three proinflammatory cytokines, IL-1beta, TNF-alpha, and IFN-gamma, are driven by two separable mechanisms. NOS-2 expression requires a burst of p38 MAPK activity, while the activation of the resulting enzyme protein requires MEK/ERK-dependent BH4 (tetrahydrobiopterin) synthesis between 24 and 24.5 h after adding the cytokines to the culture medium. Here we show that NOS-2 expression in the activated astrocytes requires that the culture medium contain 1.8 mM Ca2+, but it is unaffected by inhibiting calcium-sensing receptors (CASRs) with NPS 89636. However, NOS-2 activation is inhibited by NPS 89626 during the MEK/ERK-dependent stage between 24 and 24.5 h after adding the cytokines, and this inhibition can be overridden by exogenous BH4. Therefore, NOS-2 expression and the subsequent BH4-dependent NOS-2-activation in human astrocytes need 1.8 mM Ca2+ to be in the culture medium, while NOS-2 activation also needs functional CASRs between 24 and 24.5 h after cytokine addition. These findings raise the possibility that calcilytic drugs prevent NO-induced damage and death of human neurons.

Identification of calcium sensing receptor (CaSR) mRNA-expressing cells in normal and injured rat brain

“…In the present work, CaSR expression in the oligodendrocytes is in agreement with previous data (Ferry et al, 2000), whereas the absence of CaSR in astrocytes and microglia is not (Chattopadhyay et al, 1999. It is noteworthy that although in an earlier study failed to detect expression of the CaSR in primary culture of astrocytes isolated from rat brain, later these and others authors demonstrated the presence of CaSR mRNA and protein in primary human astrocytes Dal Pra et al, 2005). The function of the CaSR in normal oligodendroglial cells is currently unknown, but it is reasonable to hypothesize a role in sensing and regulating local ionic homeostasy during neuronal activity.…”

Mechanisms of interleukin-1β-induced Ca2+ signals in mouse cortical astrocytes: roles of store- and receptor-operated Ca2+ entry

“…Disruption of Ca 2ϩ homeostasis might be one of the principal mechanisms by which IL-1␤ manifests its neurotoxicity. Indeed, there have been several reports (11,36) showing that Ca 2ϩ disregulation plays an important role in the production of IL-1␤ and nitric oxide synthase in cytokine-stimulated astrocytes.…”

Novel Role of the Calcium-Sensing Receptor in Blood Pressure Modulation

“…55 The CaR was shown previously to stimulate NO production in other cells. 56,57 The CaR was also found in VSMCs from the gerbilline spiral modiolar artery, 58 from rat aorta, 7 and from human aorta. 59 However, it should be noted that some investigators have been unable to demonstrate the presence of the CaR in aortic VSMCs.…”