Expression of α₂-adrenergic receptor subtypes in prenatal rat spinal cord

The results of molecular cloning have revealed three subtypes of the α₂-adrenergic receptors (α₂ AR) that have been defined α₂C10 (α_2A), α₂C2 (α_2B) and α₂C4 (α_2C). The differential expression of α₂ AR subtypes is affected by developmental factors in rat submandibular gland, lung and brain. In the spinal cord of postnatal and adult rats, α_2A and α_2C AR subtypes are expressed and appear to mediate pain perception. However, the relative expression of α₂ AR subtypes in the prenatal spinal cord is unknown. In the present study subtype-specific antibodies and reverse transcription-polymerase chain reaction (RT-PCR) were used to determine the expression and localization of the α₂ AR subtypes in sections of embryonic day 14 rat spinal cords and primary cultures of cells isolated from these cords. Spinal cords were removed from day 14 embryos, and were sectioned or used for the preparation of cell cultures. After 9 days in culture, neurons were examined by immunofluorescence microscopy or used for preparation of total RNA. In both intact spinal cords and isolated cells, positive immunoreactivity was detected with antibodies against α_2A and α_2B subtypes, but not with antibodies against the α_2C subtype. Using a dual-labeling approach, anti-α_2A and anti-α_2B immunoreactivity was present on the same population of neurons. RT-PCR results were consistent with immunofluorescence studies, and showed that mRNA encoding the α_2A and α_2B subtypes was present in total RNA prepared from primary cultures of rat spinal cord neurons. In contrast to spinal cords of postnatal or adult rats that express α_2A and α_2C AR subtypes on different neurons, prenatal spinal cords contain α_2A and α_2B AR subtypes, and these two subtypes appear to be co-expressed in the same cells.