Regulation of Volume-Sensitive Osmolyte Efflux from Human SH-SY5Y Neuroblastoma Cells following Activation of Lysophospholipid Receptors

Abstract

The ability of the lysophospholipids sphingosine 1-phosphate (S1P) and lysophosphatidic acid (LPA) to promote the release of the organic osmolyte taurine in response to hypoosmotic stress has been examined. Incubation of SH-SY5Y neuroblastoma cells under hypoosmotic conditions (230 mOsM) resulted in a time-dependent release of taurine that was markedly enhanced (3-7-fold) by the addition of micromolar concentrations of either S1P or LPA. At optimal concentrations, the effects of S1P and LPA on taurine efflux were additive and mediated via distinct receptors. Inclusion of 1,9-dideoxyfoskolin, 5-nitro-2-(3-phenylpropylamino benzoic acid, or 4-[(2-butyl-6,7-dicloro-2-cyclopentyl-2,3-dihydro-1-oxo-1H-inden-5-yl)oxy]-butanoic acid blocked the ability of both lysophospholipids to enhance taurine release, indicating the mediation of a volume-sensitive organic osmolyte and anion channel. Both S1P and LPA elicited robust increases in intracellular calcium concentration that were attenuated by the removal of extracellular calcium, abolished by the depletion of intracellular calcium with thapsigargin, and were independent of phosphoinositide turnover. Taurine efflux mediated by S1P and LPA was unaffected by the removal of extracellular calcium but was attenuated by depletion of intracellular calcium (34 -38%) and by inhibition of protein kinase C (PKC) with chelerythrine (38 -72%). When intracellular calcium was depleted and PKC was inhibited, S1P-or LPAstimulated taurine efflux was inhibited by 80%. Pretreatment of the cells with pertussis toxin, toxin B, or cytochalasin D had no effect on lysophospholipid-stimulated taurine efflux. The results indicate that both S1P and LPA receptors facilitate osmolyte release via a phospholipase C-independent mechanism that requires the availability of intracellular calcium and PKC activity.

Impact of SLC6A Transporters in Physiological Taurine Transport at the Blood–Retinal Barrier and in the Liver

“…42) On the other hand, in response to hypotonic stress, osmolyte extrusion recovers cell volume, and studies in TR-iBRB2 cells suggests that the volume of the retinal capillary endothelial cells is regulated by volume-sensitive organic osmolyte and anion channel (VSOAC) that releases taurine in response to hypoosmotic stress. 41,43) Subsequently, studies under hypotonic conditions suggested the induction of taurine transport of the retinal capillary endothelial cells in the presence of sphingosine-1-phosphate (S1P), a ligand of G proteincoupled receptors (GPCRs), 41) and the induction of volumesensitive taurine release by retinal capillary endothelial cells through GPCRs is also consistent with a previous report on GPCRs associated with osmolarity regulation. 44) In the retina, nerve activity is constant, and the various types of neurotransmitters released cause osmolarity fluctuations in the retinal microenvironment, such as the retinal extracellular fluid (ECF).…”

Two conventional protein kinase C isoforms, α and βI, are involved in the ATP‐induced activation of volume‐regulated anion channel and glutamate release in cultured astrocytes

“…This is in agreement with a number of previous findings (Du and Sorota 1999;Falktoft and Lambert 2004;Mongin and Kimelberg 2005;Heacock et al 2006;Cheema et al 2007). Additional work is needed to establish if aforementioned PKC dependence of hypoosmotic VRAC activation in HeLa, HTC and Mz-ChA-1 cells (Roman et al 1998;Hermoso et al 2004) is restricted to a few cell types or is more universal phenomenon.…”

“…However, a number of laboratories have demonstrated that stimulation of GPCRs, including several metabotropic P2Y receptors for ATP and ADP, produces limited activation of a VRAC-like pathway in non-swollen cells and greatly increases its activity in cells exposed to hypoosmotic medium (Jeremic et al 2001;Mongin and Kimelberg 2002;Mongin and Kimelberg 2003;Loveday et al 2003;Darby et al 2003;Franco et al 2004). Depending on the experimental system and the GPCR involved, PKCs have been found to play major or minor roles in the regulation of VRAC activity by GPCR agonists (Loveday et al 2003;Falktoft and Lambert 2004;Mongin and Kimelberg 2005;Heacock et al 2006;Cheema et al 2007;Ramos-Mandujano et al 2007). In HeLa cells, HTC rat hepatoma cells, and Mz-ChA-1 human cholangiocarcinoma cells, PKC activity was found to be critical for hypotonic (i.e.…”

“…Several groups discovered that ATP and agonists of several other G-protein coupled receptors (GPCR) strongly increase VRAC activity in swollen cells. GPCR agonists may also produce limited VRAC activation in non-swollen cells of both neural and non-neural origin (Tilly et al 1994;Mongin and Kimelberg 2002;Loveday et al 2003;Darby et al 2003;Franco et al 2004;Falktoft and Lambert 2004;Heacock et al 2006;Cheema et al 2007). Regulation of VRAC activity by several purinergic (ATP) receptors may be especially important in the brain.…”

Muscarinic Receptor Stimulation of d-Aspartate Uptake into Human SH-SY5Y Neuroblastoma Cells Is Attenuated by Hypoosmolarity

“…In C 6 glioma cells, activation of several phosphoinositide-linked receptors has previously been demonstrated to facilitate D-aspartate uptake (Najimi et al, 2002(Najimi et al, , 2005Fournier et al, 2004). In this context, it may be relevant that of the GPCRs tested in SH-SY5Y neuroblastoma only the mAChR couples to phosphoinositide hydrolysis in this cell line (Heacock et al, 2006). D-Aspartate uptake into SH-SY5Y cells was mediated via a high-affinity, NaCl-dependent, saturable transport system that could be blocked by L␤BA, a potent inhibitor of the neuron-specific glutamate transporter EAAT3.…”