Characteristics of P2X7-like receptor activated by adenosine triphosphate in HIT-T15 cells

Dong Hyeon Lee, Eun Gi Kim, Kyusang Park, Seong Woo Jeong, Indeok Kong, Joong Woo Lee

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

OBJECTIVES: The study examined the presence of a P2X7 receptor subtype and its functional roles in pancreatic β cells. METHODS: In a hamster β-cell line, HIT-T15 cells, purinergic stimulation was investigated using fluorometry, electrophysiology, flow cytometry, and electrophoresis. RESULTS: Adenosine triphosphate (ATP) and 2′-3′-O-(4-benzoylbenzoyl)adenosine 5′-triphosphate (BzATP) increased in the intracellular free Ca concentration, with an EC50 of 398.0 and 136.6 μM, respectively. Preincubation with oxidized ATP, a P2X7 receptor antagonist, inhibited the ATP- and BzATP-induced increase in the intracellular Ca level. The BzATP-induced increase in the intracellular Ca level was dependent on the extracellular Ca concentration. The extracellular Mg had a significant effect on the ATP-induced increase in the intracellular Ca level. The ATP also induced depolarization like high potassium chloride. In the voltage-clamp experiments, ATP evoked inward currents, which were reversed at almost 0 mV. The ATP stimulated the slow influx of ethidium bromide, indicating permeability to larger molecules. Flow cytometry showed that the number of hypodiploid cells (A0), which are indicative of apoptosis, increased when the cells were exposed to ATP for 24 hours. The ATP also induced DNA fragmentation. CONCLUSIONS: These results suggest that the HIT-T15 cells have endogenous P2X7-like receptors and that purinergic stimulation increased the level of intracellular Ca, depolarization, inward current, permeability, and apoptosis.

Original languageEnglish
Pages (from-to)53-62
Number of pages10
JournalPancreas
Volume35
Issue number1
DOIs
Publication statusPublished - 2007 Jul 1

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Purinergic P2X7 Receptors
Adenosine Triphosphate
Permeability
Flow Cytometry
Apoptosis
Fluorometry
Ethidium
Potassium Chloride
Electrophysiology
DNA Fragmentation
Cricetinae
Electrophoresis

All Science Journal Classification (ASJC) codes

  • Internal Medicine
  • Endocrinology, Diabetes and Metabolism
  • Hepatology
  • Endocrinology

Cite this

Lee, Dong Hyeon ; Kim, Eun Gi ; Park, Kyusang ; Jeong, Seong Woo ; Kong, Indeok ; Lee, Joong Woo. / Characteristics of P2X7-like receptor activated by adenosine triphosphate in HIT-T15 cells. In: Pancreas. 2007 ; Vol. 35, No. 1. pp. 53-62.
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Characteristics of P2X7-like receptor activated by adenosine triphosphate in HIT-T15 cells. / Lee, Dong Hyeon; Kim, Eun Gi; Park, Kyusang; Jeong, Seong Woo; Kong, Indeok; Lee, Joong Woo.

In: Pancreas, Vol. 35, No. 1, 01.07.2007, p. 53-62.

Research output: Contribution to journalArticle

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AU - Lee, Dong Hyeon

AU - Kim, Eun Gi

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N2 - OBJECTIVES: The study examined the presence of a P2X7 receptor subtype and its functional roles in pancreatic β cells. METHODS: In a hamster β-cell line, HIT-T15 cells, purinergic stimulation was investigated using fluorometry, electrophysiology, flow cytometry, and electrophoresis. RESULTS: Adenosine triphosphate (ATP) and 2′-3′-O-(4-benzoylbenzoyl)adenosine 5′-triphosphate (BzATP) increased in the intracellular free Ca concentration, with an EC50 of 398.0 and 136.6 μM, respectively. Preincubation with oxidized ATP, a P2X7 receptor antagonist, inhibited the ATP- and BzATP-induced increase in the intracellular Ca level. The BzATP-induced increase in the intracellular Ca level was dependent on the extracellular Ca concentration. The extracellular Mg had a significant effect on the ATP-induced increase in the intracellular Ca level. The ATP also induced depolarization like high potassium chloride. In the voltage-clamp experiments, ATP evoked inward currents, which were reversed at almost 0 mV. The ATP stimulated the slow influx of ethidium bromide, indicating permeability to larger molecules. Flow cytometry showed that the number of hypodiploid cells (A0), which are indicative of apoptosis, increased when the cells were exposed to ATP for 24 hours. The ATP also induced DNA fragmentation. CONCLUSIONS: These results suggest that the HIT-T15 cells have endogenous P2X7-like receptors and that purinergic stimulation increased the level of intracellular Ca, depolarization, inward current, permeability, and apoptosis.

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