Naslov
Identification and characterization of a novel population of P2X4 receptors in hepatocytes

Autori
Emmett, Daniel ; Feranchak, Andrew ; Kilic, Gordan ; Puljak, Livia ; Miller, Bonnie ; McWilliams, Brian ; Doctor, Brian ; Fitz, J. Gregory

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Izvornik
Hepatology / - , 2005

Skup
Liver Meeting

Mjesto i datum
San Francisco (CA), Sjedinjene Američke Države, 11.11.2005. - 15.11.2005

Vrsta sudjelovanja
Poster

Vrsta recenzije
Međunarodna recenzija

Ključne riječi
P2X4 receptors; hepatocytes

Sažetak
In intact liver, extracellular ATP acts as a potent signaling molecule that stimulates glucose release and increases intracellular Ca2+ concentration. The specific receptors involved are not known. Recently, a novel class of P2X receptors has been identified in neurons and other cell types. These function as receptorgated cation channels, where agonist binding (ATP, Zn2+) leads to opening of a nonselective cation pore permeable to both Na+ and Ca2+. Accordingly, the purpose of these studies was to evaluate whether P2X receptors are expressed by hepatocytes and contribute to ATP-dependent calcium signaling. Methods: Studies were performed in isolated rat hepatocytes and HTC rat hepatoma cells. Membrane ion permeability was measured using whole-cell patch clamp techniques, and intracellular Ca2 concentration was assessed using fluo-3 fluorescence. Results: Transcripts for three P2X isoforms (P2X3, -4, and -7) were detectable by RT-PCR ; and both P2X4 and P2X7 proteins were detectable by Western blotting. A full length cDNA encoding P2X4 was identified from HTC cells. The sequence was identical to rat brain P2X4 receptor, and encoded a protein of 64 kDa with two putative transmembrane domains. The receptor appears to be present in both the basolateral and canalicular domains as detected by i) Western blotting of isolated hepatocyte subcellular fractions and by ii)immunohistochemistry staining of intact rat liver. However, receptor density was consistently greater in the canalicular domain. To assess whether these receptors are functional, whole cell patch clamp studies were performed. Exposure to a P2X4 receptor agonist, BzATP, caused a rapid increase in membrane conductance. Currents were carried predominately by influx of Na+, activated instantaneously, exhibited a peak current density of -27 +/- 3 pA/pF (-80 mV), and then decreased to a lower sustained plateau. In parallel studies, BzATP caused a rapid increase in intracellular [Ca2+]. The peak response decreased from 153 +/- 17 nM to 37 +/- 1 nM (p<0.01) in the presence versus absence of extracellular calcium. Similar results were obtained by exposure to Zn2+ (10 uM), which also functions as a P2X4 receptor agonist. Summary: These studies provide both molecular and functional evidence that P2X4 receptors are expressed by hepatocytes and mediate ATP-dependent Na and Ca2 influx. The observations that these receptors are localized to both the basolateral and canalicular domains, and are regulated by divalent cations, suggest multiple modes of action. These studies also demonstrate that P2X4 receptors may play an important role in the autocrine regulation of liver function.

Izvorni jezik
Engleski

Znanstvena područja
Temeljne medicinske znanosti

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