The relationship between agonist states of the M1 muscarinic receptor and the hydrolysis of inositol lipids in transfected murine fibroblast cells (B82) expressing different receptor densities

L. Mei, J. Lai, H. I. Yamamura, W. R. Roeske

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Abstract

We studied the relationship between the M1 muscarinic receptor density and the receptor-mediated hydrolysis of inositol lipids in cloned murine fibroblast B82 cells which were transfected with the M1 muscarinic receptor gene. Of the seven clones examined, the M1 muscarinic receptor densities in these cells characterized by (-)[3H]methyl-3-quinuclidinyl benzilate [(-)-[3H]MQNB binding ranged from 12 fmol/106 cells in LK3-1 cells to 260 fmol/106 cells in the LK3-8 cells. Carbachol/(-)[3H]MQNB competition curves for the LK3-1 cells (with low receptor density) had a Hill coefficient close to unity. The competition curves foir carbachol in the clones with higher receptor densities had Hill coefficients less than 1 and were best fitted by a computerized nonlinear least-squares regression program for the two-site model. The percentage of the M1 muscarinic receptors which had high affinity for carbachol decreased as the receptor density increased, suggesting that the presence of endogenous factors in these cells may be important for the agonist affinity state of the receptor. Concentration-response curves for carbachol-stimulated [3H]inositol monophosphate ([3H]IP1) accumulation were also obtained. A significant correlation was observed between the density of M1 muscarinic receptor with high affinity for carbachol and the maximum [3H]IP1 accumulation in these cells. There is no significant difference among the EC50 values and the dissociation constant of high-affinity state values of the carbachol/(-)[3H]MQNB competition curves. These results suggest that the high-affinity state for carbachol may be the functional state of the M1 muscarinic receptors in these transfected B82 cells. Although a linear correlation between the total M1 muscarinic receptor density and the maximum [3H]IP1 accumulation was observed, there was evidence for the existence of spare M1 receptors in the clones with high receptor densities but not in those with low receptor densities.

Original languageEnglish (US)
Pages (from-to)90-97
Number of pages8
JournalJournal of Pharmacology and Experimental Therapeutics
Volume251
Issue number1
StatePublished - Jan 1 1989

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Muscarinic M1 Receptors
Inositol
Carbachol
Hydrolysis
Fibroblasts
Lipids
Clone Cells
Quinuclidinyl Benzilate
Least-Squares Analysis
Cell Count

ASJC Scopus subject areas

  • Pharmacology

Cite this

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title = "The relationship between agonist states of the M1 muscarinic receptor and the hydrolysis of inositol lipids in transfected murine fibroblast cells (B82) expressing different receptor densities",
abstract = "We studied the relationship between the M1 muscarinic receptor density and the receptor-mediated hydrolysis of inositol lipids in cloned murine fibroblast B82 cells which were transfected with the M1 muscarinic receptor gene. Of the seven clones examined, the M1 muscarinic receptor densities in these cells characterized by (-)[3H]methyl-3-quinuclidinyl benzilate [(-)-[3H]MQNB binding ranged from 12 fmol/106 cells in LK3-1 cells to 260 fmol/106 cells in the LK3-8 cells. Carbachol/(-)[3H]MQNB competition curves for the LK3-1 cells (with low receptor density) had a Hill coefficient close to unity. The competition curves foir carbachol in the clones with higher receptor densities had Hill coefficients less than 1 and were best fitted by a computerized nonlinear least-squares regression program for the two-site model. The percentage of the M1 muscarinic receptors which had high affinity for carbachol decreased as the receptor density increased, suggesting that the presence of endogenous factors in these cells may be important for the agonist affinity state of the receptor. Concentration-response curves for carbachol-stimulated [3H]inositol monophosphate ([3H]IP1) accumulation were also obtained. A significant correlation was observed between the density of M1 muscarinic receptor with high affinity for carbachol and the maximum [3H]IP1 accumulation in these cells. There is no significant difference among the EC50 values and the dissociation constant of high-affinity state values of the carbachol/(-)[3H]MQNB competition curves. These results suggest that the high-affinity state for carbachol may be the functional state of the M1 muscarinic receptors in these transfected B82 cells. Although a linear correlation between the total M1 muscarinic receptor density and the maximum [3H]IP1 accumulation was observed, there was evidence for the existence of spare M1 receptors in the clones with high receptor densities but not in those with low receptor densities.",
author = "L. Mei and J. Lai and Yamamura, {H. I.} and Roeske, {W. R.}",
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T1 - The relationship between agonist states of the M1 muscarinic receptor and the hydrolysis of inositol lipids in transfected murine fibroblast cells (B82) expressing different receptor densities

AU - Mei, L.

AU - Lai, J.

AU - Yamamura, H. I.

AU - Roeske, W. R.

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N2 - We studied the relationship between the M1 muscarinic receptor density and the receptor-mediated hydrolysis of inositol lipids in cloned murine fibroblast B82 cells which were transfected with the M1 muscarinic receptor gene. Of the seven clones examined, the M1 muscarinic receptor densities in these cells characterized by (-)[3H]methyl-3-quinuclidinyl benzilate [(-)-[3H]MQNB binding ranged from 12 fmol/106 cells in LK3-1 cells to 260 fmol/106 cells in the LK3-8 cells. Carbachol/(-)[3H]MQNB competition curves for the LK3-1 cells (with low receptor density) had a Hill coefficient close to unity. The competition curves foir carbachol in the clones with higher receptor densities had Hill coefficients less than 1 and were best fitted by a computerized nonlinear least-squares regression program for the two-site model. The percentage of the M1 muscarinic receptors which had high affinity for carbachol decreased as the receptor density increased, suggesting that the presence of endogenous factors in these cells may be important for the agonist affinity state of the receptor. Concentration-response curves for carbachol-stimulated [3H]inositol monophosphate ([3H]IP1) accumulation were also obtained. A significant correlation was observed between the density of M1 muscarinic receptor with high affinity for carbachol and the maximum [3H]IP1 accumulation in these cells. There is no significant difference among the EC50 values and the dissociation constant of high-affinity state values of the carbachol/(-)[3H]MQNB competition curves. These results suggest that the high-affinity state for carbachol may be the functional state of the M1 muscarinic receptors in these transfected B82 cells. Although a linear correlation between the total M1 muscarinic receptor density and the maximum [3H]IP1 accumulation was observed, there was evidence for the existence of spare M1 receptors in the clones with high receptor densities but not in those with low receptor densities.

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