Identification of alpha 1C-adrenergic receptor mRNA in bovine retinal pigment epithelium.
OBJECTIVE
To examine the localization of a novel alpha 1-adrenergic receptor subtype of alpha 1C receptor in the eye, we compared the amount of alpha 1C receptor transcripts in bovine retinal pigment epithelium (RPE) and in neural retina by employing reverse transcription of mRNA and the polymerase chain reaction (RT-PCR) assay.


METHODS
RT-PCR assay with specific primers for the alpha 1C-adrenergic receptor and for beta-actin showed linear relationships between input quantity of RNA and the amount of amplified PCR products for the alpha 1C-adrenergic receptor and also for beta-actin, when PCR was conducted for 24 and 15 cycles, respectively.


RESULTS
The RT-PCR assay demonstrated that the spontaneous expression level of the alpha 1C-adrenergic receptor mRNA was much higher in bovine RPE than in neural retina; the alpha 1C-adrenergic receptor/beta-actin ratio from RPE was 0.33 +/- 0.15 (n = 4), whereas that from neural retina was virtually zero.


CONCLUSIONS
The RT-PCR assay is a sensitive semiquantitative method for a low abundance mRNA in a limited number of cells. Using the alpha 1C receptor as a model, we demonstrated the usefulness of this assay by showing the uneven distribution of the alpha 1C receptor transcripts in bovine RPE cells and neural retina.
Department of Pediatric Pharmacology, National Children's Medical Research Center, Tokyo, Japan.
K
K
Horie
A
A
Hirasawa
K
K
Masuda
G
G
Tsujimoto
D000199:Actins / Q000032:analysis / Q000235:genetics; D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D002417:Cattle; D004587:Electrophoresis, Agar Gel; D005786:Gene Expression Regulation; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D010857:Pigment Epithelium of Eye / Q000737:chemistry*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis*; D011942:Receptors, Adrenergic, alpha / Q000032:analysis* / Q000235:genetics; D012160:Retina / Q000737:chemistry*
D000199:Actins; D015345:Oligonucleotide Probes; D012333:RNA, Messenger; D011942:Receptors, Adrenergic, alpha