Multiple functional P2X and P2Y receptors in the luminal and basolateral membranes of pancreatic duct cells

Xiang Luo, Weizhong Zheng, Ming Yan, Min Goo Lee, Shmuel Muallem

Research output: Contribution to journalArticle

64 Citations (Scopus)

Abstract

Purinergic receptors in the basolateral and luminal membranes of the pancreatic duct can act by a feedback mechanism to coordinate transport activity in the two membranes during ductal secretion. The goal of the present work was to identify and localize the functional P2 receptors (P2R) in the rat pancreatic duct. The lack of selective agonists and/or antagonists for any of the cloned P2R dictated the use of molecular and functional approaches to the characterization of ductal P2R. For the molecular studies, RNA was prepared from microdissected pancreatic intralobular ducts and was shown to be free of mRNA for amylase and endothelial nitric oxide synthase (markers for acinar and endothelial cells, respectively). A new procedure is described to obtain an enriched preparation of single duct cells suitable for electrophysiological studies. Localization of P2R was achieved by testing the effect of various P2R agonists on intracellular Ca2+ concentration ([Ca2+](i)) of microperfused intralobular ducts. RT-PCR analysis suggested the expression of six subtypes of P2R in the pancreatic duct: three P2YR and three P2XR. Activation of Cl- current by various nucleotides and coupling of the receptors activated by these nucleotides to G proteins confirmed the expression of multiple P2R in duct cells. Measurement of [Ca2+](i) in microperfused intralobular ducts suggested the expression of P2X1R, P2X4R, probably P2X7R, and as yet unidentified P2YR, possibly P2Y1R, in the basolateral membrane. Expression of P2Y2R, P2Y4R, and P2X7R was found in the luminal membrane. The unprecedented expression of such a variety of P2R in one cell type, many capable of activating Cl- channels, suggests that these receptors may have an important role in pancreatic duct cell function.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Cell Physiology
Volume277
Issue number2 46-2
StatePublished - 1999

Fingerprint

Pancreatic Ducts
Ducts
Membranes
Nucleotides
Purinergic Receptors
Acinar Cells
Nitric Oxide Synthase Type III
Amylases
GTP-Binding Proteins
Endothelial Cells
RNA
Polymerase Chain Reaction
Messenger RNA
Endothelial cells
Rats
Chemical activation
Feedback
Testing

Keywords

  • Chloride channels
  • Intracellular calcium
  • Purinergic receptors
  • Rat pancreatic duct

ASJC Scopus subject areas

  • Clinical Biochemistry
  • Cell Biology
  • Physiology
  • Physiology (medical)

Cite this

Multiple functional P2X and P2Y receptors in the luminal and basolateral membranes of pancreatic duct cells. / Luo, Xiang; Zheng, Weizhong; Yan, Ming; Lee, Min Goo; Muallem, Shmuel.

In: American Journal of Physiology - Cell Physiology, Vol. 277, No. 2 46-2, 1999.

Research output: Contribution to journalArticle

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AU - Muallem, Shmuel

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N2 - Purinergic receptors in the basolateral and luminal membranes of the pancreatic duct can act by a feedback mechanism to coordinate transport activity in the two membranes during ductal secretion. The goal of the present work was to identify and localize the functional P2 receptors (P2R) in the rat pancreatic duct. The lack of selective agonists and/or antagonists for any of the cloned P2R dictated the use of molecular and functional approaches to the characterization of ductal P2R. For the molecular studies, RNA was prepared from microdissected pancreatic intralobular ducts and was shown to be free of mRNA for amylase and endothelial nitric oxide synthase (markers for acinar and endothelial cells, respectively). A new procedure is described to obtain an enriched preparation of single duct cells suitable for electrophysiological studies. Localization of P2R was achieved by testing the effect of various P2R agonists on intracellular Ca2+ concentration ([Ca2+](i)) of microperfused intralobular ducts. RT-PCR analysis suggested the expression of six subtypes of P2R in the pancreatic duct: three P2YR and three P2XR. Activation of Cl- current by various nucleotides and coupling of the receptors activated by these nucleotides to G proteins confirmed the expression of multiple P2R in duct cells. Measurement of [Ca2+](i) in microperfused intralobular ducts suggested the expression of P2X1R, P2X4R, probably P2X7R, and as yet unidentified P2YR, possibly P2Y1R, in the basolateral membrane. Expression of P2Y2R, P2Y4R, and P2X7R was found in the luminal membrane. The unprecedented expression of such a variety of P2R in one cell type, many capable of activating Cl- channels, suggests that these receptors may have an important role in pancreatic duct cell function.

AB - Purinergic receptors in the basolateral and luminal membranes of the pancreatic duct can act by a feedback mechanism to coordinate transport activity in the two membranes during ductal secretion. The goal of the present work was to identify and localize the functional P2 receptors (P2R) in the rat pancreatic duct. The lack of selective agonists and/or antagonists for any of the cloned P2R dictated the use of molecular and functional approaches to the characterization of ductal P2R. For the molecular studies, RNA was prepared from microdissected pancreatic intralobular ducts and was shown to be free of mRNA for amylase and endothelial nitric oxide synthase (markers for acinar and endothelial cells, respectively). A new procedure is described to obtain an enriched preparation of single duct cells suitable for electrophysiological studies. Localization of P2R was achieved by testing the effect of various P2R agonists on intracellular Ca2+ concentration ([Ca2+](i)) of microperfused intralobular ducts. RT-PCR analysis suggested the expression of six subtypes of P2R in the pancreatic duct: three P2YR and three P2XR. Activation of Cl- current by various nucleotides and coupling of the receptors activated by these nucleotides to G proteins confirmed the expression of multiple P2R in duct cells. Measurement of [Ca2+](i) in microperfused intralobular ducts suggested the expression of P2X1R, P2X4R, probably P2X7R, and as yet unidentified P2YR, possibly P2Y1R, in the basolateral membrane. Expression of P2Y2R, P2Y4R, and P2X7R was found in the luminal membrane. The unprecedented expression of such a variety of P2R in one cell type, many capable of activating Cl- channels, suggests that these receptors may have an important role in pancreatic duct cell function.

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