Yonsei Med J.  2011 May;52(3):420-428. 10.3349/ymj.2011.52.3.420.

C-Jun NH2-Terminal Kinase Contributes to Dexmedetomidine-Induced Contraction in Isolated Rat Aortic Smooth Muscle

• Ok SH
• Jeong YS
• Kim JG
• Lee SM
• Sung HJ
• Kim HJ
• Chang KC
• Kwon SC
• Sohn JT

• KMID: 1777013
• DOI: http://doi.org/10.3349/ymj.2011.52.3.420

Abstract

PURPOSE
Dexmedetomidine, a full agonist of alpha2B-adrenoceptors, is used for analgesia and sedation in the intensive care units. Dexmedetomidine produces an initial transient hypertension due to the activation of post-junctional alpha2B-adrenoceptors on vascular smooth muscle cells (SMCs). The aims of this in vitro study were to identify mitogen-activated protein kinase (MAPK) isoforms that are primarily involved in full, alpha2B-adrenoceptor agonist, dexmedetomidine-induced contraction of isolated rat aortic SMCs.
MATERIALS AND METHODS
Rat thoracic aortic rings without endothelium were isolated and suspended for isometric tension recording. Cumulative dexmedetomidine (10(-9) to 10(-6) M) dose-response curves were generated in the presence or absence of extracellular signal-regulated kinase (ERK) inhibitor PD 98059, p38 MAPK inhibitor SB 203580, c-Jun NH2-terminal kinase (JNK) inhibitor SP 600125, L-type calcium channel blocker (verapamil and nifedipine), and alpha2-adrenoceptor inhibitor atipamezole. Dexmedetomidine-induced phosphorylation of ERK, JNK, and p38 MAPK in rat aortic SMCs was detected using Western blotting.
RESULTS
SP 600125 (10(-6) to 10(-5) M) attenuated dexmedetomidine-evoked contraction in a concentration-dependent manner, whereas PD 98059 had no effect on dexmedetomidine-induced contraction. SB 203580 (10(-5) M) attenuated dexmedetomidine-induced contraction. Dexmedetomidine-evoked contractions were both abolished by atipamezole and attenuated by verapamil and nifedipine. Dexmedetomidine induced phosphorylation of JNK and p38 MAPK in rat aortic SMCs, but did not induce phosphorylation of ERK.
CONCLUSION
Dexmedetomidine-induced contraction involves a JNK- and p38 MAPK-mediated pathway downstream of alpha2-adrenoceptor stimulation in rat aortic SMCs. In addition, dexmedetomidine-induced contractions are primarily dependent on calcium influx via L-type calcium channels.

Keyword

Dexmedetomidine; mitogen-activated protein kinase; alpha2B-adrenoceptors; hypertension; rat aorta

MeSH Terms

Adrenergic alpha-2 Receptor Agonists/*pharmacology
Animals
Anthracenes/pharmacology
Aorta/cytology
Dexmedetomidine/*pharmacology
Enzyme Inhibitors/pharmacology
Extracellular Signal-Regulated MAP Kinases/antagonists & inhibitors/physiology
Flavonoids/pharmacology
Imidazoles/pharmacology
JNK Mitogen-Activated Protein Kinases/antagonists & inhibitors/*physiology
Male
*Muscle Contraction
Muscle, Smooth, Vascular/drug effects/enzymology/*physiology
Protein Isoforms/antagonists & inhibitors/physiology
Pyridines/pharmacology
Rats
Rats, Sprague-Dawley
p38 Mitogen-Activated Protein Kinases/antagonists & inhibitors/physiology

Figure

• 

  Fig. 1 Medetomidine enantiomers concentration-response curves in endothelium-denuded rings. Data represent the mean±SD and are expressed as the percentage of the maximal contraction induced by isotonic 30 mM KCl (isotonic 30 mM KCl-induced contraction: 100%=2.30±0.60 g [n=8] and 100%=1.90±0.20 g [n=8] for endothelium-denuded rings with dexmedetomidine and levomedetomidine, respectively). *p<0.001 compared with 10-9 M dexmedetomidine. Maximal contraction: †p<0.001 compared with levomedetomidine.

• 

  Fig. 2 Effects of PD 98059 (10-5 M), SB 203580 (10-6, 10-5 M), and SP 600125 (10-6, 3×10-6, and 10-5 M) on dexmedetomidine concentration-response curves in endothelium-denuded rings. All data represent the mean±SD and are expressed as the percentage of the maximal contraction induced by isotonic 30 mM KCl. (A) Isotonic 30 mM KCl-induced contraction: 100%=2.42±0.63 g (n=6) and 100%=2.51±0.21 g (n=6) for endothelium-denuded rings with no drug and 10-5 M PD 98059-pretreated endothelium-denuded rings, respectively. (B) Isotonic 30 mM KCl-induced contraction: 100%=2.57±0.29 g (n=7), 100%=2.81±0.30 g (n=6), and 100%=2.21±0.52 g (n=7) for endothelium-denuded rings with no drug and SB 203580 (10-6 M)- and (10-5 M)-pretreated endothelium-denuded rings, respectively. Maximal contraction: *p<0.001 compared with no drug. (C) Isotonic 30 mM KCl-induced contraction: 100%=2.50±0.42 g (n=7), 100%=2.43±0.42 g (n=6), 100%=2.34±0.29 g (n=7) and 100%=2.50±0.31 g (n=5) for endothelium-denuded rings with no drug and SP 600125 (10-6 M)-, (3×10-6 M)-, and (10-5 M)-pretreated endothelium-denuded rings, respectively. Maximal contraction: *p<0.001 compared with no drug, †p<0.001 compared with 10-6 M SP 600125, ‡p<0.001 compared with 3×10-6 M SP 600125.

• 

  Fig. 3 (A) Effect of dexmedetomidine (DMT, 3×10-8, 10-7, 3×10-7, 10-6 M) on 30 mM potassium chloride (KCl)-induced contraction in endothelium-denuded aorta. Data represent the mean±SD and are expressed as the relative percentage of the maximal contraction induced by initial 30 mM KCl (100%=2.75±0.48 g [n=6], 100%=2.71±0.78 g [n=6], 100%=2.83±0.47 g [n=6], 100%=2.68±0.45 g [n=6] and 100%=2.41±0.28 g [n=5] for endothelium-denuded aorta with no drug, 3×10-8 DMT, 10-7 DMT, 3×10-7 DMT and 10-6 M DMT, respectively). *p=0.002, †p=0.001, ‡p=0.002 and §p=0.006 compared with no drug. (B) Effects of verapamil and atipamezole on dexmedetomidine concentration- response curves in endothelium-denuded rings. Data represent the mean±SD and are expressed as the percentage of the maximal contraction induced by isotonic 30 mM KCl (isotonic 30 mM KCl-induced contraction: 100%=2.37±0.53 g [n=6], 100%=2.64±0.59 g [n=5] and 100%=1.47±0.24 g [n=5] for endothelium-denuded rings with no drug, the verapamil (10-5 M)- and atipamezole (10-4 M)-pretreated endothelium-denuded rings, respectively). Maximal contraction: *p<0.001 compared with no drug. (C) Effects of nifedipine on dexmedetomidine concentration-response curves in endothelium-denuded rings. Data represent the mean±SD and are expressed as the percentage of the maximal contraction induced by isotonic 30 mM KCl (isotonic 30 mM KCl-induced contraction: 100%=2.44±0.51 g [n=6] and 100%=2.45±0.48 g [n=6] for endothelium-denuded rings with no drug and the nifedipine (5×10-6 M)-pretreated endothelium-denuded rings, respectively). Maximal contraction: *p<0.001 compared with no drug.

• 

  Fig. 4 Effect of dexmedetomidine (DMT) on the activation of exracellular signal-regulated kinase (ERK), p38 mitogen-activated protein kinase (MAPK), and c-Jun NH2-terminal kinase (JNK) in rat aortic vascular smooth muscle cells (VSMCs). VSMCs were treated with DMT (10-7 or 10-6 M) in a time-dependent manner (5, 10, 30, or 60 min). (A) Phosphorylation of ERK, p38 MAPK, and JNK was examined by Western blot analysis as described in the methods. (B) Band intensities at 10 min after 10-6 M dexmedetomidine pretreatment were assessed by scanning densitometry. Data are presented as the mean±SD of three independent experiments. *p=0.039 and †p=0.045 compared with control.

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