RESEARCH

Calcium acts as an intracellular (second) messenger to regulate a diverse array of cellular functions, from cell division and differentiation to cell death. The generation and control of intracellular calcium signals depends on the dynamic relationships among extracellular, cytosolic, and intraorganelle calcium concentrations. For cells, the extracellular medium represents an unlimited and tightly controlled reservoir of calcium. Many intracellular organelles, including the endoplasmic/sarcoplasmic reticulum, Golgi network, mitochondria, endosomes, lysosomes, and secretory vesicles, also contain high concentrations of calcium, which allow them to serve as an additional, but limited, reservoir. As in extracellular medium, the total calcium concentration in intracellular organelles is in the millimolar dose-range. In contrast, the cytosol contains only 10-20% of the total intracellular calcium, much of which is bound to soluble cytosolic proteins and membranes, making it inaccessible to many calcium-controlled cellular processes. In unstimulated cells, the intracellular free calcium concentration is between 50 and 200 nM, whereas in stimulated cells it can increase to micromolar concentrations.

Over one hundred neurotransmitters and hormones, acting on their respective receptors and receptor-channels, relay their signals by altering intracellular calcium. For the generation of calcium signals, cells depend on the large electrochemical gradient of calcium across the plasma membrane (termed calcium influx), or across the endoplasmic reticulum membrane (termed calcium mobilization). There are several pathways controlling calcium influx. Ligand-gated receptor-channels, including nicotinic actetyl choline receptor-channels, glutamate receptor-channels, and purinergic receptor-channels, require the binding of an extracellular ligand, such as neurotransmitters, to open the channel pore. Many of these channels are permeable to calcium, but also conduct other ions. In contrast, voltage-gated calcium channels are highly selective for calcium and are opened by a decrease in the electrical potential across the plasma membrane, which is termed membrane depolarization. Many G protein-coupled receptors stimulate voltage-gated calcium influx by activating depolarizing channels or inhibiting hyperpolarizing channels. Intracellular calcium sources also provide a mechanism for the rapid propagation of calcium signals within the cell, which is important in the control of intraorganelle function. Calcium mobilization occurs in many non-excitable and excitable cell types, is activated by G protein-coupled and tyrosine kinase receptors, and is mediated through inositol (1,4,5)-trisphosphate)-sensitive receptor-channels that are associated with the endoplasmic/sarcoplasmic reticulum. Many excitable cells also express ryanodine-sensitive calcium release channels in their endoplasmic/sarcoplasmic reticulum, and these are coupled to voltage-gated calcium channels by a mechanism termed calcium-induced calcium release.

The main objective of current investigations in the Section on Cellular Signaling is to clarify the mechanisms that control calcium signaling and to eludicate the role of calcium ions as intracellular messenger in neuroendocrine and endocrine cells. The approach in the laboratory has been to characterize calcium signaling in these cells from a biophysical, physiological, and pharmacological
point of view. More recently, we have begun to use molecular biological approaches to identify molecules participating in the generation and control of calcium signals, as well as to address the issues of how the calcium signaling function in these cells is related to the structure of individual receptors and channels. Current research includes studies of the receptor- and intracellular
messenger-mediated control of voltage-gated calcium influx; the structure and function of purinergic receptor-channels; cyclic nucleotide-controlled membrane excitability and calcium signaling; the role of calcium in controlling plasma membrane and endoplasmic reticulum excitability and the relevance of the pattern of calcium signaling to cell-type specific hormone secretion patterns. We believe that the results already gained from these projects and those of other investigators have indicated the
enormous complexity and physiological relevance of the calcium signaling system in endocrine cells, as well as the need for future investigations. The information we are currently obtaining, as well as our projected research, should continue to be useful to investigators and clinicians in the endocrine field, as well as to the broad scientific community.

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SECTION CHIEF

Stanko S. Stojilovic received a Ph.D. in Physiology/Neuroendocrinology from the University of Novi Sad, Serbia, in 1982. He was Assistant Professor of Physiology at the Institute of Biology, University of Novi Sad, before joining NICHD as a Guest Researcher in 1985, where he worked in the laboratory of Kevin J. Catt. This was followed by a tenure-track appointment in 1993 and tenure appointment in 1998 in the Endocrinology and Reproduction Research Branch of NICHD. He currently serves as Head of the Section on Cellular Signaling.

Stanko S. Stojilkovic, Ph.D.
Endocrinology and Reproduction Research Branch
NICHD, Building 49, Room 6A36
Bethesda, MD 20892-4510
USA

Telephone: 301-496-1638
Fax: 301-594-7031
Email: stankos@helix.nih.gov

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Personnel:

· Silvana A Andric, Ph.D., Bldg. 49; Rm. 6C-28; tel: 301-496-8732; email: andrics@mail.nih.gov
· Mu-Lan He, Ph.D., Bldg. 49; Rm. 6C-28; tel: 301-496-7783; email: hemu@mail.nih.gov
· Arturo E. Gonzales Iglesias, Ph.D., Bldg. 49, Rm 6C-20; tel: 301-496-8164; email iglesiaas@mail.nih.gov
· Tatjana S. Kostic, Ph.D., Bldg. 49; Rm. 6C-28; tel: 301-496-7783; email: kostict@mail.nih.gov
· Melanija Tomic, Ph.D., Bldg. 49, Rm. 6C20; tel: 301-496-8164; email: melatom@box-m.nih.gov

Collaborators:

· Arthur Sherman, Ph.D., Chief, Mathematical Research Branch, NIDDK; web site: http://mrb.niddk.nih.gov/
· Olaf Ortmann, M.D., Ph.D., Professor, Department of Obstetrics and Gynecology, University of Lubeck, Germany, D-23538
· Gozoh Tsujimoto, M.D., Ph.D., Director, Department of Molecular, Cell Pharmacology; National Children's Medical Research Center 3-35-31, Taishi-do, Setagaya-Ku, Tokyo 154, Japan; http://pharmac.nch.go.jp/
· Radmila Z Kovacevic, Ph.D., Chief, Department of Physiology, Institute of Biology, Faculty of Sciences, 2 Dositeja Obradovica Square, 21000 Novi Sad, Serbia, Yugoslavia; email: radmilak@unsim.ns.ac.yu

Former Postdoctoral Fellows:

· Mirjana Cesnjaj, M.D., Ph.D., Loyola University Medical Center, Maywood, IL 60153
· Taka-aki Koshimizu, M.D., Ph.D., Second Department of Internal Medicine, Yamanashi Medical University, Tamaho-chyo 1110, Yamanashi 409-3898, Japan
· Agnieszka Lachowicz, M.D., Ph.D., Department of Experimental Endocrinology and Hormone Diagnostics, Medical University of Lodz, Lodz, Poland
· Lixin Zheng, M.D., Shady Grove Adventist Hospital, Rockville, MD 20850
· Fredrick Van Goor, Ph.D., Aurora Biosciences Corp., San Diego, CA 92121
· Dragoslava Zivadinovic, Ph.D., University of Texas, Galveston, TX 77555

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BIBLIOGRAPHY

Van Goor F, Li YX, Stojilkovic SS. (2001) Paradoxical role of large-conductance calcium-activated K+ (BK) channels in controlling action potential-driven Ca2+ entry in anterior pituitary cells. J Neurosci. 21(16):5902-15. pdf

Van Goor F, Zivadinovic D, Martinez-Fuentes AJ, Stojilkovic SS. (2001) Dependence of pituitary hormone secretion on the pattern of spontaneous voltage-gated calcium influx. Cell type-specific action potential secretion coupling. J Biol Chem. 276(36):33840-6. pdf

Van Goor F, Zivadinovic D, Stojilkovic SS. (2001) Differential expression of ionic channels in rat anterior pituitary cells. Mol Endocrinol. 15(7):1222-36. pdf

Stojilkovic SS, Koshimizu T. (2001) Signaling by extracellular nucleotides in anterior pituitary cells. Trends Endocrinol Metab. 12(5):218-25. pdf

Kostic TS, Andric SA, Stojilkovic SS. (2001) Spontaneous and receptor-controlled soluble guanylyl cyclase activity in anterior pituitary cells. Mol Endocrinol. 15(6):1010-22. pdf

Andric SA, Kostic TS, Tomic M, Koshimizu TA, Stojilkovic SS. (2001) Dependence of Soluble Guanylyl Cyclase Activity on Calcium Signaling in Pituitary Cells. J. Biol Chem. 276(1):844-849. pdf

LeBeau AP, Van Goor F , Stojilkovic SS, Sherman A. (2000) Modeling of membrane excitability in gonadotropin-releasing hormone-secreting hypothalamic neurons regulated by Ca2+-mobilizing and adenylyl cyclase-coupled receptors. J Neurosci. 20(24):9290-7. pdf

Koshimizu TA, Tomic M, Wong AO, Zivadinovic D, Stojilkovic SS. (2000) Characterization of purinergic receptors and receptor-channels expressed in anterior pituitary cells. Endocrinology. 141(11):4091-9. pdf

Koshimizu TA, Van Goor F, Tomic M, Wong AO, Tanoue A, Tsujimoto G, Stojilkovic SS. (2000) Characterization of calcium signaling by purinergic receptor-channels expressed in excitable cells. Mol Pharmacol. 58(5):936-45. pdf

Van Goor F, LeBeau AP, Krsmanovic LZ, Sherman A, Catt KJ, Stojilkovic SS. (2000) Amplitude-dependent spike-broadening and enhanced Ca(2+) signaling in GnRH-secreting neurons. Biophys J. 79(3):1310-23. pdf

Koshimizu T, Koshimizu M, Stojilkovic SS. (1999) Contributions of the C-terminal domain to the control of P2X receptor desensitization. J Biol Chem. 274(53):37651-7. pdf

Tomic M, Koshimizu T, Yuan D, Andric SA, Zivadinovic D, Stojilkovic SS. (1999) Characterization of a plasma membrane calcium oscillator in rat pituitary somatotrophs. J Biol Chem. 274(50):35693-702. pdf

Tomic M, Zivadinovic D, Van Goor F, Yuan D, Koshimizu T, Stojilkovic SS. (1999) Expression of Ca(2+)-mobilizing endothelin(A) receptors and their role in the control of Ca(2+) influx and growth hormone secretion in pituitary somatotrophs. J. Neurosci. 19(18):7721-31. pdf

Van Goor F, Krsmanovic LZ, Catt KJ, Stojilkovic SS. (1999) Control of action potential-driven calcium influx in GT1 neurons by the activation status of sodium and calcium channels . Mol. Endocrinol. 13(4):587-603. pdf

Van Goor F, Krsmanovic LZ, Catt KJ, Stojilkovic SS. (1999) Coordinate regulation of gonadotropin-releasing hormone neuronal firing patterns by cytosolic calcium and store depletion. Proc Natl Acad Sci U S A. 96(7):4101-6. pdf

Katzur AC, Koshimizu T, Tomic M, Schultze-Mosgau A, Ortmann O, Stojilkovic SS. (1999) Expression and responsiveness of P2Y2 receptors in human endometrial cancer cell lines. J Clin Endocrinol Metab. 84(11):4085-91. pdf

Koshimizu T, Tomic M, Van Goor F, Stojilkovic SS. (1998) Functional role of alternative splicing in pituitary P2X2 receptor-channel activation and desensitization. Mol Endocrinol. 12(7):901-13. pdf

Koshimizu T, Tomic M, Koshimizu M, Stojilkovic SS. (1997) Identification of amino acid residues contributing to desensitization of the P2X2 receptor channel. J. Biol Chem. 273(21):12853-7. pdf

Lachowicz A, Van Goor F, Katzur AC, Bonhomme G, Stojilkovic SS. (1997) Uncoupling of calcium mobilization and entry pathways in endothelin-stimulated pituitary lactotrophs. J. Biol Chem. 272(45):28308-14. pdf

Ortmann O, Tomic M, Weiss JM, Diedrich K, Stojilkovic SS. (1997) Dual action of androgen on calcium signaling and luteinizing hormone secretion in pituitary gonadotrophs. Cell Calcium. 24(3):223-31. pdf

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