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Guanine nucleotide regulatory protein alterations in the Milan hypertensive rat strain
C J Clark1, G Milligan, J M Connell
1MRC Blood Pressure Unit, Western Infirmary, Glasgow, UK.
Objective:
To examine whether the altered regulation of adenylyl cyclase that has been reported in vascular tissues from spontaneously hypertensive rats is also evident in the Milan hypertensive (MHS) rat strain.
Design:
The plasma membranes of vascular smooth muscle cells derived from thoracic aortae from adult (60-day-old) MHS and Milan normotensive (MNS) strain rats were studied.
Methods:
Guanine nucleotide regulatory protein (G-protein) function was inferred from adenylyl cyclase activity studies, and levels of G-protein subunits were assessed by immunoblotting. beta-Adrenergic receptor number and affinity were measured from the binding of the antagonist [125I]-cyanopindolol.
Results:
Basal adenylyl cyclase activity was increased significantly in MHS rat cell membranes, and stimulation by 0.1 mmol/l isoproterenol and 0.01 mmol/l prostaglandin E1 was significantly greater in MHS than in MNS rat cell membranes. Forskolin (at 0.1 mmol/l) resulted in a significantly greater stimulatory response in MHS membranes, which was eliminated by 0.01 mol/l NaF. Biphasic effects of GTP on isoproterenol-stimulated membranes demonstrated similar Gi function in MHS and MNS rat cell membranes, although a greater stimulatory GTP response was observed in MHS rat cell membranes. The levels of Gs alpha (both forms), Gi3 alpha and the beta-subunit were reduced in MHS rat cell membranes, whereas the levels of Gi2 alpha and Gq alpha and G11 alpha were unchanged. The number of beta-adrenoceptors was increased significantly in MHS rat cell membranes, whereas receptor affinity for the antagonist was unaltered.
Conclusions:
There are differences in adenylyl cyclase stimulatory responses in MHS rat vascular smooth muscle cell membranes. We have found evidence of reduced levels of particular G-protein subunits, altered beta-adrenoceptor-Gs coupling and increased beta-adrenoceptor number.
Insights
Milan hypertensive rats show altered adenylyl cyclase activity and G-protein regulation in vascular smooth muscle cells. These findings suggest differences in beta-adrenergic receptor signaling pathways contributing to hypertension.
Area of Science:
- Cardiovascular Physiology
- Molecular Biology
- Hypertension Research
Background:
- Altered adenylyl cyclase regulation is observed in spontaneously hypertensive rat vascular tissues.
- Understanding these alterations in other hypertensive models is crucial.
Purpose of the Study:
- To investigate adenylyl cyclase regulation in vascular smooth muscle cells of Milan hypertensive rats (MHS).
- To compare G-protein function and beta-adrenergic receptor characteristics between MHS and normotensive (MNS) rats.
Main Methods:
- Studied plasma membranes from thoracic aortae of adult MHS and MNS rats.
- Assessed adenylyl cyclase activity, G-protein subunit levels (immunoblotting), and beta-adrenergic receptor binding.
Main Results:
- Increased basal and stimulated adenylyl cyclase activity in MHS rat membranes.
- Reduced levels of specific G-protein subunits (Gs alpha, Gi3 alpha, beta-subunit) in MHS rats.
- Increased beta-adrenoceptor number without altered affinity in MHS rats.
Conclusions:
- Milan hypertensive rat vascular smooth muscle exhibits distinct adenylyl cyclase stimulatory responses.
- Evidence suggests reduced G-protein subunits, altered beta-adrenoceptor-Gs coupling, and increased beta-adrenoceptor number in MHS rats.