Hepatocytes from alpha1B-adrenoceptor knockout mice reveal compensatory adrenoceptor subtype substitution.
1 Alpha1-adrenoceptors (ARs) play an important functional role in the liver; yet little is known about their cellular location. We identified the subtypes present in wild-type (WT) and alpha1B-AR knockout (KO) mice livers at 3 and 4 months of age, and investigated their distribution in hepatocytes. 2 The fluorescent alpha1-AR antagonist quinazolinyl piperazine borate-dipyrromethene (QAPB) was used to visualise hepatic alpha1-ARs and radioligand binding with [3H]-prazosin was used to quantify the alpha1-AR population. 3 QAPB and [3H]-prazosin bound specifically to hepatic alpha1-ARs with nanomolar affinity. The cellular distribution of alpha1-ARs was similar in WT and alpha1B-AR KO hepatocytes; QAPB binding was distributed diffusely throughout the cell with no binding evident on the plasma membrane. Radioligand binding produced Bmax values as follows: 3-month WT - 76+/-3.3 fmol mg(-1); 4-month WT - 50+/-3.1 fmol mg(-1); 3-month alpha1B-AR KO - 7.4+/-0.73 fmol mg(-1); 4-month alpha1B-AR KO - 30+/-2.0 fmol mg(-1). 4 In 3- and 4-month WT liver, all antagonists acted competitively. RS100329 (alpha1A-selective) and BMY7378 (alpha1D-selective) bound with low affinities, indicating the presence of alpha1B-ARs. In 4-month alpha1B-AR KO liver prazosin produced a biphasic curve, whereas RS100329 and BMY7378 produced monophasic curves of high and low affinity, respectively, indicating the presence of alpha1A-ARs. 5 In conclusion, we have made the novel observation that alpha1-ARs can compensate for one another in the absence of the endogenously expressed receptor; yet there appears to be no subtype-specific subcellular location of alpha1-ARs; the WT livers express alpha1B-ARs, while alpha1B-AR KO livers express alpha1A-ARs. This study provides new insights into both hepatocyte and alpha1-AR biology.
Deighan
Clare
C
Autonomic Physiology Unit, Division of Neuroscience and Biomedical Systems, Institute of Biomedical and Life Sciences, University of Glasgow, Glasgow, G12-8QQ. c.deighan@bio.gla.ac.uk
Woollhead
Alison M
AM
Colston
Janet F
JF
McGrath
John C
JC
D058668:Adrenergic alpha-1 Receptor Antagonists; D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects; D001896:Boron Compounds / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D005838:Genotype; D022781:Hepatocytes / Q000166:cytology / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008815:Mice, Inbred Strains; D018345:Mice, Knockout; D018613:Microscopy, Confocal; D010879:Piperazines / Q000378:metabolism / Q000494:pharmacology; D011224:Prazosin / Q000031:analogs & derivatives / Q000378:metabolism; D011869:Radioligand Assay; D018340:Receptors, Adrenergic, alpha-1 / Q000235:genetics / Q000378:metabolism*; D013941:Thymine / Q000378:metabolism / Q000494:pharmacology; D014316:Tritium
C547714:Adra1a protein, mouse; C547717:Adra1b protein, mouse; C547720:Adra1d protein, mouse; D058668:Adrenergic alpha-1 Receptor Antagonists; D001896:Boron Compounds; D010879:Piperazines; C120711:RS 100329; D018340:Receptors, Adrenergic, alpha-1; D014316:Tritium; C053428:BMY 7378; D013941:Thymine; D011224:Prazosin