Abstract
Human SK-N-SH neuroblastoma cells expressed sigma-1 and sigma-2 receptors with similar pharmacological profiles to those of rodent-derived tissues, although sigma-2 receptors exhibited some affinity differences that might suggest heterogeneity or species differences. Structurally diverse sigma ligands produced two types of increases in intracellular (cytosolic) Ca2+ concentration ([Ca2+]i) in these cells. CB-64D, CB-64L, JL-II-147, BD737, LR172, BD1008, haloperidol, reduced haloperidol, and ibogaine all produced an immediate, dose-dependent, and transient rise in [Ca2+]i. Sigma-inactive compounds structurally similar to the most active sigma ligands and ligands for several neurotransmitter receptors produced little or no effect. The high activity of CB-64D and ibogaine (sigma-2-selective ligands) compared with the low activity of (+)-pentazocine and other (+)-benzomorphans (sigma-1-selective ligands), in addition to enantioselectivity for CB-64D over CB-64L, strongly indicated mediation by sigma-2 receptors. The effect of CB-64D and BD737 was blocked by the sigma antagonists BD1047 and BD1063, further confirming specificity as a receptor-mediated event. The transient rise in [Ca2+]i occurred in the absence of extracellular Ca2+ and was completely eliminated by pretreatment of cells with thapsigargin. Thus, sigma-2 receptors stimulate a transient release of Ca2+ from the endoplasmic reticulum. Prolonged exposure of cells to sigma-receptor ligands resulted in a latent and sustained rise in [Ca2+]i, with a pharmacological profile identical to that of the transient rise. This sustained rise in [Ca2+]i was affected by neither the removal of extracellular Ca2+ nor thapsigargin pretreatment, suggesting latent sigma-2 receptor-induced release from thapsigargin-insensitive intracellular Ca2+ stores. Sigma-2 receptors may use Ca2+ signals in producing cellular effects.
Footnotes
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Send reprint requests to: Wayne D. Bowen, Ph.D., Chief, Unit on Receptor Biochemistry and Pharmacology, Laboratory of Medicinal Chemistry, NIDDK/National Institutes of Health, Bldg. 8, Room B1-23, 8 Center Dr. MSC 0815, Bethesda, MD 20892-0815. E-mail:bowenw{at}bdg8.niddk.nih.gov
- Abbreviations:
- DTG
- 1,3-di-o-tolylguanidine
- [Ca2+]i
- intracellular (cytosolic) Ca2+ concentration
- DMEM
- Dulbecco's modified Eagle's medium
- SERCA
- sarcoplasmic-endoplasmic reticulum Ca2+-ATPase
- DPBS
- Dulbecco's phosphate-buffered saline
- IP3
- inositol-1,4,5-trisphosphate
- Red HAL
- reduced haloperidol
- DAMGO
- [d-Ala2,N-Me-Phe4,Gly-ol5]-enkephalin
- NMDA
- N-methyl-d-aspartate
- Received May 21, 1999.
- Accepted November 23, 1999.
- U.S. Government
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