Abstract
This study aimed to determine the mechanism(s) by which 1,4-dihydropyridine Ca2+ channel blockers (DHPs) enhance the binding of neurotensin (NT) to prostate cancer PC3 cells and inhibit NT-induced inositol phosphate formation. Earlier work indicated that these effects, which involved the G protein-coupled NT receptor NTR1, were indirect and required cellular metabolism or architecture. At the micromolar concentrations used, DHPs can block voltage-sensitive and store-operated Ca2+ channels, K+ channels, and Na+ channels, and can inhibit lipid peroxidation. By varying [Ca2+] and testing the effects of stimulators and inhibitors of Ca2+ influx and internal Ca2+ release, we determined that although DHPs may have inhibited inositol phosphate formation partly by blocking Ca2+ influx, the effect on NT binding was Ca2+-independent. By varying [K+] and [Na+], we showed that these ions did not contribute to either effect. For a series of DHPs, the activity order for effects on NTR1 function followed that for antioxidant ability. Antioxidant polyphenols (luteolin and resveratrol) mimicked the effects of DHPs and showed structural similarity to DHPs. Antioxidants with equal redox ability, but without structural similarity to DHPs (such as α-tocopherol, riboflavin, and N-acetyl-cysteine) were without effect. A flavoprotein oxidase inhibitor (diphenylene iodonium) and a hydroxy radical scavenger (butylated hydroxy anisole) also displayed the effects of DHPs. In conclusion, DHPs indirectly alter NTR1 function in live cells by a mechanism that depends on the drug's ability to donate hydrogen but does not simply involve sulfhydryl reduction.
Footnotes
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This work was supported by Department of Defense Grant DAMD17-00-1-0528 and National Institutes of Health Center Grant DK32520, although the opinions expressed are not necessarily those of the Department of Defense and the National Institutes of Health.
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Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
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DOI: 10.1124/jpet.103.060442.
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ABBREVIATIONS: NT, neurotensin; NTR1, neurotensin receptor subtype 1; PLC, phosphatidylinositol-specific phospholipase C; IP, inositol phosphate; CCB, Ca2+ channel blocker; VGCC, voltage-gated Ca2+ channel; DHP, 1,4-dihydropyridine; SOCC, store-operated Ca2+ channel; DTT, dithiothreitol; BAPTA-AM, 1,2-bis(o-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid tetra(acetoxymethyl) ester; BSA, bovine serum albumin; ROS, reactive oxygen species; DPI, diphenylene iodonium; BHA, butylated hydroxy anisole; SKF-96365, 1-{β-[3-(4-methoxyphenyl)propoxy]-4-methoxyphenethyl}-1H-imidazole; FPL-64176, 2,5-dimethyl-4-[2-(phenylmethyl)benzoyl]-1H-pyrrole-3-carboxylic acid methyl ester; (-)-BayK-8644, S(-)-1,4-dihydro-2,6-dimethyl-5-nitro-4[2-(trifluoromethyl)phenyl]-3-pyridine-carboxylic acid methyl ester.
- Received September 24, 2003.
- Accepted December 16, 2003.
- The American Society for Pharmacology and Experimental Therapeutics
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