Abstract
The molecular and pharmacological characteristics of muscarinic receptor subtypes in the rat parotid acinar cell line, PAR-C5, were determined and compared with native rat parotid glands to evaluate the PAR-C5 cell line as a model to study receptor-mediated secretion. Reverse transcription-polymerase chain reaction (RT-PCR) identified mRNAs for M3, M4, and M5 receptor subtypes in both PAR-C5 cells and parotid glands. Specific [N-methyl-3H]scopolamine binding in PAR-C5 and parotid membranes was to a single class of sites with mean KD values of 0.38 and 0.64 nM, respectively. Binding affinities (KI values) of muscarinic receptor subtype-selective drugs were obtained in side-by-side experiments comparing PAR-C5 cells with parotid glands. Nonlinear regression analysis indicated that competition binding curves for drugs in PAR-C5 cells and parotid glands fit best to a one-site binding model. KI values (nM) in PAR-C5 cells and parotid glands, respectively, for atropine (1.0, 2.1), darifenacin (1.2, 2.0), 4-diphenylacetoxy-N-methylpiperidine methiodide (4-DAMP) (2.9, 2.4), tripitramine (220, 180), pirenzepine (320, 720), and methoctramine (1400, 1700) were consistent with their known affinities at the M3 receptor subtype. Affinities (KB values) of muscarinic receptor subtype-selective drugs for blocking methacholine-stimulated Ca2+ mobilization were determined to show which subtype mediates Ca2+-dependent secretion in Fura-2-loaded PAR-C5 cells. KB values (nM) for atropine (0.44), 4-DAMP (0.38), zepine (140), and methoctramine (320) for blocking Ca2+ responses correlated well with their known affinities at the M3 receptor (r2 = 0.99). These results show that at the level of mRNA, receptor protein and function, PAR-C5 cells and parotid glands are similar, establishing PAR-C5 cells as an important model for muscarinic receptor-mediated secretion.
| Original language | English |
|---|---|
| Pages (from-to) | 718-726 |
| Number of pages | 9 |
| Journal | Journal of Pharmacology and Experimental Therapeutics |
| Volume | 297 |
| Issue number | 2 |
| State | Published - 2001 |
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All Science Journal Classification (ASJC) codes
- Pharmacology
Cite this
Molecular and pharmacological characterization of muscarinic receptor subtypes in a rat parotid gland cell line : Comparison with native parotid gland. / Bockman, Charles; Bradley, Michael E.; Dang, Herbert K.; Zeng, Wanyun; Scofield, Margaret A.; Dowd, Frank J.
In: Journal of Pharmacology and Experimental Therapeutics, Vol. 297, No. 2, 2001, p. 718-726.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Molecular and pharmacological characterization of muscarinic receptor subtypes in a rat parotid gland cell line
T2 - Comparison with native parotid gland
AU - Bockman, Charles
AU - Bradley, Michael E.
AU - Dang, Herbert K.
AU - Zeng, Wanyun
AU - Scofield, Margaret A.
AU - Dowd, Frank J.
PY - 2001
Y1 - 2001
N2 - The molecular and pharmacological characteristics of muscarinic receptor subtypes in the rat parotid acinar cell line, PAR-C5, were determined and compared with native rat parotid glands to evaluate the PAR-C5 cell line as a model to study receptor-mediated secretion. Reverse transcription-polymerase chain reaction (RT-PCR) identified mRNAs for M3, M4, and M5 receptor subtypes in both PAR-C5 cells and parotid glands. Specific [N-methyl-3H]scopolamine binding in PAR-C5 and parotid membranes was to a single class of sites with mean KD values of 0.38 and 0.64 nM, respectively. Binding affinities (KI values) of muscarinic receptor subtype-selective drugs were obtained in side-by-side experiments comparing PAR-C5 cells with parotid glands. Nonlinear regression analysis indicated that competition binding curves for drugs in PAR-C5 cells and parotid glands fit best to a one-site binding model. KI values (nM) in PAR-C5 cells and parotid glands, respectively, for atropine (1.0, 2.1), darifenacin (1.2, 2.0), 4-diphenylacetoxy-N-methylpiperidine methiodide (4-DAMP) (2.9, 2.4), tripitramine (220, 180), pirenzepine (320, 720), and methoctramine (1400, 1700) were consistent with their known affinities at the M3 receptor subtype. Affinities (KB values) of muscarinic receptor subtype-selective drugs for blocking methacholine-stimulated Ca2+ mobilization were determined to show which subtype mediates Ca2+-dependent secretion in Fura-2-loaded PAR-C5 cells. KB values (nM) for atropine (0.44), 4-DAMP (0.38), zepine (140), and methoctramine (320) for blocking Ca2+ responses correlated well with their known affinities at the M3 receptor (r2 = 0.99). These results show that at the level of mRNA, receptor protein and function, PAR-C5 cells and parotid glands are similar, establishing PAR-C5 cells as an important model for muscarinic receptor-mediated secretion.
AB - The molecular and pharmacological characteristics of muscarinic receptor subtypes in the rat parotid acinar cell line, PAR-C5, were determined and compared with native rat parotid glands to evaluate the PAR-C5 cell line as a model to study receptor-mediated secretion. Reverse transcription-polymerase chain reaction (RT-PCR) identified mRNAs for M3, M4, and M5 receptor subtypes in both PAR-C5 cells and parotid glands. Specific [N-methyl-3H]scopolamine binding in PAR-C5 and parotid membranes was to a single class of sites with mean KD values of 0.38 and 0.64 nM, respectively. Binding affinities (KI values) of muscarinic receptor subtype-selective drugs were obtained in side-by-side experiments comparing PAR-C5 cells with parotid glands. Nonlinear regression analysis indicated that competition binding curves for drugs in PAR-C5 cells and parotid glands fit best to a one-site binding model. KI values (nM) in PAR-C5 cells and parotid glands, respectively, for atropine (1.0, 2.1), darifenacin (1.2, 2.0), 4-diphenylacetoxy-N-methylpiperidine methiodide (4-DAMP) (2.9, 2.4), tripitramine (220, 180), pirenzepine (320, 720), and methoctramine (1400, 1700) were consistent with their known affinities at the M3 receptor subtype. Affinities (KB values) of muscarinic receptor subtype-selective drugs for blocking methacholine-stimulated Ca2+ mobilization were determined to show which subtype mediates Ca2+-dependent secretion in Fura-2-loaded PAR-C5 cells. KB values (nM) for atropine (0.44), 4-DAMP (0.38), zepine (140), and methoctramine (320) for blocking Ca2+ responses correlated well with their known affinities at the M3 receptor (r2 = 0.99). These results show that at the level of mRNA, receptor protein and function, PAR-C5 cells and parotid glands are similar, establishing PAR-C5 cells as an important model for muscarinic receptor-mediated secretion.
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M3 - Article
VL - 297
SP - 718
EP - 726
JO - Journal of Pharmacology and Experimental Therapeutics
JF - Journal of Pharmacology and Experimental Therapeutics
SN - 0022-3565
IS - 2
ER -