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Identification of store-independent and store-operated Ca2+ conductances in Caenorhabditis elegans intestinal epithelial cells.
J Gen Physiol. 2003 Aug; 122(2):207-23.JG

Abstract

The nematode Caenorhabditis elegans offers significant experimental advantages for defining the genetic basis of diverse biological processes. Genetic and physiological analyses have demonstrated that inositol-1,4,5-trisphosphate (IP3)-dependent Ca2+ oscillations in intestinal epithelial cells play a central role in regulating the nematode defecation cycle, an ultradian rhythm with a periodicity of 45-50 s. Patch clamp studies combined with behavioral assays and forward and reverse genetic screening would provide a powerful approach for defining the molecular details of oscillatory Ca2+ signaling. However, electrophysiological characterization of the intestinal epithelium has not been possible because of its relative inaccessibility. We developed primary intestinal epithelial cell cultures that circumvent this problem. Intestinal cells express two highly Ca2+-selective, voltage-independent conductances. One conductance, IORCa, is constitutively active, exhibits strong outward rectification, is 60-70-fold more selective for Ca2+ than Na+, is inhibited by intracellular Mg2+ with a K1/2 of 692 microM, and is insensitive to Ca2+ store depletion. Inhibition of IORCa with high intracellular Mg2+ concentrations revealed the presence of a small amplitude conductance that was activated by passive depletion of intracellular Ca2+ stores. Active depletion of Ca2+ stores with IP3 or ionomycin increased the rate of current activation approximately 8- and approximately 22-fold compared with passive store depletion. The store-operated conductance, ISOC, exhibits strong inward rectification, and the channel is highly selective for Ca2+ over monovalent cations with a divalent cation selectivity sequence of Ca2+ > Ba2+ approximately Sr2+. Reversal potentials for ISOC could not be detected accurately between 0 and +80 mV, suggesting that PCa/PNa of the channel may exceed 1,000:1. Lanthanum, SKF 96365, and 2-APB inhibit both IORCa and ISOC reversibly. Our studies provide the first detailed electrophysiological characterization of voltage-independent Ca2+ conductances in C. elegans and form the foundation for ongoing genetic and molecular studies aimed at identifying the genes that encode the intestinal cell channels, for defining mechanisms of channel regulation and for defining their roles in oscillatory Ca2+ signaling.

Authors+Show Affiliations

Department of Anesthesiology, Vanderbilt University Medical Center, Nashville, TN 37232, USA.No affiliation info availableNo affiliation info available

Pub Type(s)

Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

12860924

Citation

Estevez, Ana Y., et al. "Identification of Store-independent and Store-operated Ca2+ Conductances in Caenorhabditis Elegans Intestinal Epithelial Cells." The Journal of General Physiology, vol. 122, no. 2, 2003, pp. 207-23.
Estevez AY, Roberts RK, Strange K. Identification of store-independent and store-operated Ca2+ conductances in Caenorhabditis elegans intestinal epithelial cells. J Gen Physiol. 2003;122(2):207-23.
Estevez, A. Y., Roberts, R. K., & Strange, K. (2003). Identification of store-independent and store-operated Ca2+ conductances in Caenorhabditis elegans intestinal epithelial cells. The Journal of General Physiology, 122(2), 207-23.
Estevez AY, Roberts RK, Strange K. Identification of Store-independent and Store-operated Ca2+ Conductances in Caenorhabditis Elegans Intestinal Epithelial Cells. J Gen Physiol. 2003;122(2):207-23. PubMed PMID: 12860924.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Identification of store-independent and store-operated Ca2+ conductances in Caenorhabditis elegans intestinal epithelial cells. AU - Estevez,Ana Y, AU - Roberts,Randolph K, AU - Strange,Kevin, Y1 - 2003/07/14/ PY - 2003/7/16/pubmed PY - 2004/5/7/medline PY - 2003/7/16/entrez SP - 207 EP - 23 JF - The Journal of general physiology JO - J Gen Physiol VL - 122 IS - 2 N2 - The nematode Caenorhabditis elegans offers significant experimental advantages for defining the genetic basis of diverse biological processes. Genetic and physiological analyses have demonstrated that inositol-1,4,5-trisphosphate (IP3)-dependent Ca2+ oscillations in intestinal epithelial cells play a central role in regulating the nematode defecation cycle, an ultradian rhythm with a periodicity of 45-50 s. Patch clamp studies combined with behavioral assays and forward and reverse genetic screening would provide a powerful approach for defining the molecular details of oscillatory Ca2+ signaling. However, electrophysiological characterization of the intestinal epithelium has not been possible because of its relative inaccessibility. We developed primary intestinal epithelial cell cultures that circumvent this problem. Intestinal cells express two highly Ca2+-selective, voltage-independent conductances. One conductance, IORCa, is constitutively active, exhibits strong outward rectification, is 60-70-fold more selective for Ca2+ than Na+, is inhibited by intracellular Mg2+ with a K1/2 of 692 microM, and is insensitive to Ca2+ store depletion. Inhibition of IORCa with high intracellular Mg2+ concentrations revealed the presence of a small amplitude conductance that was activated by passive depletion of intracellular Ca2+ stores. Active depletion of Ca2+ stores with IP3 or ionomycin increased the rate of current activation approximately 8- and approximately 22-fold compared with passive store depletion. The store-operated conductance, ISOC, exhibits strong inward rectification, and the channel is highly selective for Ca2+ over monovalent cations with a divalent cation selectivity sequence of Ca2+ > Ba2+ approximately Sr2+. Reversal potentials for ISOC could not be detected accurately between 0 and +80 mV, suggesting that PCa/PNa of the channel may exceed 1,000:1. Lanthanum, SKF 96365, and 2-APB inhibit both IORCa and ISOC reversibly. Our studies provide the first detailed electrophysiological characterization of voltage-independent Ca2+ conductances in C. elegans and form the foundation for ongoing genetic and molecular studies aimed at identifying the genes that encode the intestinal cell channels, for defining mechanisms of channel regulation and for defining their roles in oscillatory Ca2+ signaling. SN - 0022-1295 UR - https://www.unboundmedicine.com/medline/citation/12860924/Identification_of_store_independent_and_store_operated_Ca2+_conductances_in_Caenorhabditis_elegans_intestinal_epithelial_cells_ DB - PRIME DP - Unbound Medicine ER -