Quantitative analysis of mu and delta opioid receptor gene expression in rat brain and peripheral ganglia using competitive polymerase chain reaction.
Competitive polymerase chain reaction assays following reverse transcription have been developed for quantitative analysis of delta and mu opioid receptor gene expression. The assay was used to obtain quantitative measurements of mu and delta opioid receptor expression levels in different brain regions and sensory and sympathetic ganglia in the rat. The assays provide accurate estimates of the relative levels of receptor messenger RNAs by the inclusion in the assays of known amounts of internal standards with the same sequence, except for a small deletion, as the target complementary DNA. The amplification products of target and competitor can be distinguished by size, and their amounts measured by densitometry. Expression of mu and delta opioid receptor messenger RNAs in different regions of the rat brain, somatic and visceral sensory and sympathetic ganglia was investigated using this method. In the brain the highest density of delta receptor messenger RNA was detected in the olfactory bulb, followed by the striatum. The mu receptor was expressed at highest levels in the midbrain-hypothalamic region. All the sensory ganglia studied expressed both mu and delta opioid receptor messenger RNAs. In the nodose ganglion we observed the highest level of mu receptor messenger RNA of any structure studied; in the trigeminal ganglion the level was about 10 times lower than that in the nodose ganglion. Among the dorsal root ganglia, mu receptor messenger RNA density was highest in the lumbar region, followed by the thoracic and cervical regions. The sympathetic superior cervical ganglion expressed a very low level of mu message. Delta receptor messenger RNA was detected only in the sensory ganglia, at levels that were considerably lower than in the striatum. The reverse transcription polymerase chain reaction assay is quantitatively reliable for comparison of messenger RNA levels between different RNA extracts, and sensitive enough to permit the detection and assay of mu and delta opioid receptor gene expression in a single pair of sensory or autonomic ganglia from the rat.
Búzás
B
B
Department of Pharmacology, Uniformed Services University, Bethesda, MD 20814, USA.
Cox
B M
BM
D000818:Animals; D001923:Brain Chemistry / Q000502:physiology*; D004247:DNA / Q000096:biosynthesis; D005724:Ganglia / Q000378:metabolism*; D005727:Ganglia, Spinal / Q000378:metabolism; D015870:Gene Expression / Q000502:physiology*; D008297:Male; D009620:Nodose Ganglion / Q000378:metabolism; D016133:Polymerase Chain Reaction; D012313:RNA / Q000096:biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D017207:Rats, Sprague-Dawley; D017465:Receptors, Opioid, delta / Q000096:biosynthesis* / Q000235:genetics*; D017450:Receptors, Opioid, mu / Q000096:biosynthesis* / Q000235:genetics*; D017783:Superior Cervical Ganglion / Q000378:metabolism; D012668:Trigeminal Ganglion / Q000378:metabolism
D012333:RNA, Messenger; D017465:Receptors, Opioid, delta; D017450:Receptors, Opioid, mu; D012313:RNA; D004247:DNA