α1D-Adrenergic Receptors and Mitogen-Activated Protein Kinase Mediate Increased Protein Synthesis by Arterial Smooth Muscle
Abstract
Catecholamines may influence vascular smooth muscle cell (SMC) growth and vascular hypertrophic diseases. We previously demonstrated that stimulation of α1-adrenoceptors (AR) causes hypertrophy of vascular SMCs in vitro and in situ. Here, we used adult rat aorta SMCs that express α1D- and α1B-ARs (but not α1A-ARs) in vitro to examine the mechanisms and α1-AR subtypes involved. Norepinephrine (NE) increased protein synthesis and content in a time- and dose-dependent manner. To identify the responsible α1-AR subtype, we first documented the selectivity of two α1-AR subtype antagonists, BMY 7378 (α1D-AR antagonist) and chloroethylclonidine (CEC; α1B-AR antagonist), using Rat-1 fibroblasts stably transfected with the three different rodent α1-AR cDNAs. NE dose-dependently increased protein synthesis in each cell line. In α1D fibroblasts, BMY 7378 inhibited growth and protected α1D-ARs from CEC alkylation while having little blocking or protecting effect on the growth induced by stimulation of fibroblasts that express α1A- or α1B-ARs. In rat aorta SMCs, pretreatment with CEC in the presence of BMY 7378 to protect α1D-ARs had no effect on NE-induced protein synthesis. BMY 7378 inhibited the SMC growth response with a pKb of 8.4. NE caused rapid and transient p42-p44 mitogen-activated protein kinase (MAPK) activation that was α1D-AR dependent. Furthermore, NE caused tyrosine phosphorylation of multiple cellular proteins, phosphorylation of Raf-1, and stimulation of c-fos mRNA expression in aorta SMCs. The selective MAPK kinase inhibitor PD 98059 inhibited NE-induced protein synthesis and MAPK activation with IC50values of 2.3 and 1.6 μm, respectively. These data demonstrate that SMC growth induced by NE is mediated by α1D-ARs that couple to activation of the MAPK cascade.
Footnotes
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Send reprint requests to: Dr. James E. Faber, Department of Physiology, 474 MSRB, University of North Carolina, Chapel Hill, NC 27599-7545. E-mail: jefaber{at}med.unc.edu
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↵1 J. E. Faber, X. Xin, N. Yang, and A. D. Eckhart, unpublished observations.
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This study was supported by National Institutes of Health Grant HL52610.
- Abbreviations:
- SMC
- smooth muscle cell
- AR
- adrenoceptor
- FB
- fibroblast
- NE
- norepinephrine
- CEC
- chloroethylclonidine
- MAPK
- mitogen-activated protein kinase
- MEK
- mitogen-activated protein kinase kinase
- SSC
- standard saline citrate
- PBS
- phosphate-buffered saline
- TCA
- trichloroacetic acid
- bp
- base pair(s)
- GAPDH
- glyceraldehyde-3-phosphate dehydrogenase
- ANOVA
- analysis of variance
- VC
- vena cava
- EGF
- epidermal growth factor
- PKC
- protein kinase C
- RPA
- RNase protection assay
- 5-MU
- 5-methyl-urapidil
- ERK
- extracellular signal-regulated kinase
- PDGF
- platelet-derived growth factor
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- Received December 2, 1996.
- Accepted January 28, 1997.
- The American Society for Pharmacology and Experimental Therapeutics
