Activation of α2A-adrenergic receptors in human trabecular meshwork cells stimulates MAP kinase activity

W. D. Stamer, R. W. Snyder, T. H. Burkey, John W Regan

Research output: Contribution to journalArticle

Abstract

Purpose: Previously, we have demonstrated the presence of α2A-adrenergic receptors (AR) in primary cultures of human trabecular meshwork (HTM) cells by immunofluorescence microscopy and the inhibition of forskolin-stimulated adenylyl cyclase. Since it has been shown in transfected cells that the α2A-AR can activate mitogen activated protein (MAP) kinase, we were interested in the possibility that the endogenous α2A-ARs in HTM cells might be coupled to MAP kinase activity and possibly cellular proliferation. Methods: HTM cells were incubated in the presence of serum and bFGF for 48 hours and then were serum-starved for 24 hours. Cells were stimulated with either: phorbol ester myristate acetate (100nM), dexmedetomidine (DMT, 100 nM), DMT (100 nM) plus rauwolscine (10 μM) or DMT (100 nM) plus atipamezole (10 μM). Lysates were prepared and MAP kinase activity was measured by the incorporation of γ32P-ATP into myelin basic protein. Results: Activation of α2A-ARs by DMT resulted in a dose-dependent stimulation of MAP kinase activity. Preincubation of cells with α2A-AR antagonists, rauwolscine or atipamezole, significantly reduced the activation of MAP kinase by DMT. Western blot further demonstrated that the amount of immunoreactive MAP kinase was the same in both control and agonist-treated cells which is consistent with an increase in the activity, rather than the amount of MAP kinase. Conclusions: Activation of α2A-adrenergic receptors in human trabecular meshwork cells stimulates MAP kinase activity and, therefore, may be involved with the proliferation of these cells.

Original languageEnglish (US)
JournalInvestigative Ophthalmology and Visual Science
Volume37
Issue number3
StatePublished - Feb 15 1996

Fingerprint

Trabecular Meshwork
Mitogen-Activated Protein Kinases
Adrenergic Receptors
Yohimbine
Cell Proliferation
Far-Western Blotting
Dexmedetomidine
Adrenergic Antagonists
Myelin Basic Protein
Phorbol Esters
Tetradecanoylphorbol Acetate
Colforsin
Serum
Fluorescence Microscopy
Adenylyl Cyclases
Adenosine Triphosphate

ASJC Scopus subject areas

  • Ophthalmology

Cite this

Activation of α2A-adrenergic receptors in human trabecular meshwork cells stimulates MAP kinase activity. / Stamer, W. D.; Snyder, R. W.; Burkey, T. H.; Regan, John W.

In: Investigative Ophthalmology and Visual Science, Vol. 37, No. 3, 15.02.1996.

Research output: Contribution to journalArticle

@article{3d0d253b7b74468f93602ba7896c5590,
title = "Activation of α2A-adrenergic receptors in human trabecular meshwork cells stimulates MAP kinase activity",
abstract = "Purpose: Previously, we have demonstrated the presence of α2A-adrenergic receptors (AR) in primary cultures of human trabecular meshwork (HTM) cells by immunofluorescence microscopy and the inhibition of forskolin-stimulated adenylyl cyclase. Since it has been shown in transfected cells that the α2A-AR can activate mitogen activated protein (MAP) kinase, we were interested in the possibility that the endogenous α2A-ARs in HTM cells might be coupled to MAP kinase activity and possibly cellular proliferation. Methods: HTM cells were incubated in the presence of serum and bFGF for 48 hours and then were serum-starved for 24 hours. Cells were stimulated with either: phorbol ester myristate acetate (100nM), dexmedetomidine (DMT, 100 nM), DMT (100 nM) plus rauwolscine (10 μM) or DMT (100 nM) plus atipamezole (10 μM). Lysates were prepared and MAP kinase activity was measured by the incorporation of γ32P-ATP into myelin basic protein. Results: Activation of α2A-ARs by DMT resulted in a dose-dependent stimulation of MAP kinase activity. Preincubation of cells with α2A-AR antagonists, rauwolscine or atipamezole, significantly reduced the activation of MAP kinase by DMT. Western blot further demonstrated that the amount of immunoreactive MAP kinase was the same in both control and agonist-treated cells which is consistent with an increase in the activity, rather than the amount of MAP kinase. Conclusions: Activation of α2A-adrenergic receptors in human trabecular meshwork cells stimulates MAP kinase activity and, therefore, may be involved with the proliferation of these cells.",
author = "Stamer, {W. D.} and Snyder, {R. W.} and Burkey, {T. H.} and Regan, {John W}",
year = "1996",
month = "2",
day = "15",
language = "English (US)",
volume = "37",
journal = "Investigative Ophthalmology and Visual Science",
issn = "0146-0404",
publisher = "Association for Research in Vision and Ophthalmology Inc.",
number = "3",

}

TY - JOUR

T1 - Activation of α2A-adrenergic receptors in human trabecular meshwork cells stimulates MAP kinase activity

AU - Stamer, W. D.

AU - Snyder, R. W.

AU - Burkey, T. H.

AU - Regan, John W

PY - 1996/2/15

Y1 - 1996/2/15

N2 - Purpose: Previously, we have demonstrated the presence of α2A-adrenergic receptors (AR) in primary cultures of human trabecular meshwork (HTM) cells by immunofluorescence microscopy and the inhibition of forskolin-stimulated adenylyl cyclase. Since it has been shown in transfected cells that the α2A-AR can activate mitogen activated protein (MAP) kinase, we were interested in the possibility that the endogenous α2A-ARs in HTM cells might be coupled to MAP kinase activity and possibly cellular proliferation. Methods: HTM cells were incubated in the presence of serum and bFGF for 48 hours and then were serum-starved for 24 hours. Cells were stimulated with either: phorbol ester myristate acetate (100nM), dexmedetomidine (DMT, 100 nM), DMT (100 nM) plus rauwolscine (10 μM) or DMT (100 nM) plus atipamezole (10 μM). Lysates were prepared and MAP kinase activity was measured by the incorporation of γ32P-ATP into myelin basic protein. Results: Activation of α2A-ARs by DMT resulted in a dose-dependent stimulation of MAP kinase activity. Preincubation of cells with α2A-AR antagonists, rauwolscine or atipamezole, significantly reduced the activation of MAP kinase by DMT. Western blot further demonstrated that the amount of immunoreactive MAP kinase was the same in both control and agonist-treated cells which is consistent with an increase in the activity, rather than the amount of MAP kinase. Conclusions: Activation of α2A-adrenergic receptors in human trabecular meshwork cells stimulates MAP kinase activity and, therefore, may be involved with the proliferation of these cells.

AB - Purpose: Previously, we have demonstrated the presence of α2A-adrenergic receptors (AR) in primary cultures of human trabecular meshwork (HTM) cells by immunofluorescence microscopy and the inhibition of forskolin-stimulated adenylyl cyclase. Since it has been shown in transfected cells that the α2A-AR can activate mitogen activated protein (MAP) kinase, we were interested in the possibility that the endogenous α2A-ARs in HTM cells might be coupled to MAP kinase activity and possibly cellular proliferation. Methods: HTM cells were incubated in the presence of serum and bFGF for 48 hours and then were serum-starved for 24 hours. Cells were stimulated with either: phorbol ester myristate acetate (100nM), dexmedetomidine (DMT, 100 nM), DMT (100 nM) plus rauwolscine (10 μM) or DMT (100 nM) plus atipamezole (10 μM). Lysates were prepared and MAP kinase activity was measured by the incorporation of γ32P-ATP into myelin basic protein. Results: Activation of α2A-ARs by DMT resulted in a dose-dependent stimulation of MAP kinase activity. Preincubation of cells with α2A-AR antagonists, rauwolscine or atipamezole, significantly reduced the activation of MAP kinase by DMT. Western blot further demonstrated that the amount of immunoreactive MAP kinase was the same in both control and agonist-treated cells which is consistent with an increase in the activity, rather than the amount of MAP kinase. Conclusions: Activation of α2A-adrenergic receptors in human trabecular meshwork cells stimulates MAP kinase activity and, therefore, may be involved with the proliferation of these cells.

UR - http://www.scopus.com/inward/record.url?scp=33750148189&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=33750148189&partnerID=8YFLogxK

M3 - Article

AN - SCOPUS:33750148189

VL - 37

JO - Investigative Ophthalmology and Visual Science

JF - Investigative Ophthalmology and Visual Science

SN - 0146-0404

IS - 3

ER -