mRNA expression and binding sites for alpha2-adrenergic receptor subtypes in muscle layers of the ileum and spiral colon of dairy cows.
OBJECTIVE
To measure maximum binding capacity (B(max)) and levels of mRNA expression for alpha(2)-adrenergic receptor (AR) subtypes in ileal and colonic muscle layers of healthy dairy cows.


METHODS
Ileal and colonic muscle specimens from 6 freshly slaughtered cows.


METHODS
Ileal and colonic muscle layers were obtained by scraping the mucosa and submucosa from full-thickness tissue specimens. Level of mRNA expression for alpha(2)-AR subtypes was measured by real-time reverse transcriptase-PCR analysis and expressed relative to the mean mRNA expression of glyceraldehyde phosphate dehydrogenase, ubiquitin, and 18S ribosomal RNA. Binding studies were performed with tritiated RX821002 ((3)H-RX821002) and subtype-selective ligands as competitors.


RESULTS
mRNA expression for alpha(2AD)-, alpha(2B)-, and alpha(2C)-AR subtypes was similar in ileal and colonic muscle layers. The mRNA expression for alpha(2AD)-AR was significantly greater than that for alpha(2B)- and alpha(2C)-AR subtypes, representing 92%, 6%, and 2%, respectively, of the total mRNA. Binding competition of (3)H-RX821002 with BRL44408, imiloxan, and MK-912 was best fitted by a 1-site model. The B(max) of alpha(2AD)- and alpha(2C)-AR sub-types was greater than that of alpha(2B)-AR. The B(max) and level of mRNA expression were only correlated (r = 0.8) for alpha(2AD)-AR. Ratio of B(max) to mRNA expression for alpha(2C)-AR was similar to that for alpha(2B)-AR, but significantly greater than for alpha(2AD)-AR.


CONCLUSIONS
Subtypes of alpha(2)-AR in bovine intestinal muscle layers are represented by a mixture of alpha(2AD)- and alpha(2C)-ARs and of alpha(2B)-AR at a lower density. Information provided here may help in clarification of the role of AR subtypes in alpha(2)-adrenergic mechanisms regulating bovine intestinal motility.
Ontsouka
Edgar C
EC
Clinic for Ruminants, Vetsuisse Faculty of Berne, and the Division of Nutrition and Physiology, Institute of Animal Genetics, Nutrition, and Housing, Vetsuisse Faculty, University of Berne, CH-3012 Berne, Switzerland.
Blum
Jürg W
JW
Steiner
Adrian
A
Meylan
Mireille
M
D000818:Animals; D002417:Cattle / Q000235:genetics / Q000378:metabolism*; D017931:DNA Primers; D005260:Female; D019329:Idazoxan / Q000031:analogs & derivatives; D041741:Lower Gastrointestinal Tract / Q000378:metabolism*; D009130:Muscle, Smooth / Q000378:metabolism*; D011485:Protein Binding; D012333:RNA, Messenger / Q000378:metabolism*; D018341:Receptors, Adrenergic, alpha-2 / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014316:Tritium
D017931:DNA Primers; D012333:RNA, Messenger; D018341:Receptors, Adrenergic, alpha-2; D014316:Tritium; C058525:2-methoxyidazoxan; D019329:Idazoxan