GABAB receptor protein and mRNA distribution in rat spinal cord and dorsal root ganglia.
The presence of metabotropic receptors for GABA, GABAB, on primary afferent terminals in mammalian spinal cord has been previously reported. In this study we provide further evidence to support this in the rat and show that the GABAB receptor subunits GABAB1 and GABAB2 mRNA and the corresponding subunit proteins are present in the spinal cord and dorsal root ganglion. We also show that the predominant GABAB1 receptor subunit mRNA present in the afferent fibre cell body appears to be the 1a form. In frozen sections of lumbar spinal cord and dorsal root ganglia (DRG) GABAB receptors were labelled with [3H]CGP 62349 or the sections postfixed with paraformaldehyde and subjected to in situ hybridization using oligonucleotides designed to selectively hybridize with the mRNA for GABAB(1a), GABAB(1b) or GABAB2. For immunocytochemistry (ICC), sections were obtained from rats anaesthetized and perfused-fixed with paraformaldehyde. The distribution of binding sites for [3H]CGP 62349 mirrored that previously observed with [3H]GABA at GABAB sites. The density of binding sites was high in the dorsal horn but much lower in the ventral regions. By contrast, the density of mRNA (pan) was more evenly distributed across the laminae of the spinal cord. The density of mRNA detected with the pan probe was high in the DRG and distributed over the neuron cell bodies. This would accord with GABAB receptor protein being formed in the sensory neurons and transported to the primary afferent terminals. Of the GABAB1 mRNA in the DRG, approximately 90% was of the GABAB(1a) form and approximately 10% in the GABAB(1b) form. This would suggest that GABAB(1a) mRNA may be responsible for encoding presynaptic GABAB receptors on primary afferent terminals in a manner similar to that we have previously observed in the cerebellar cortex. GABAB2 mRNA was also evenly distributed across the spinal cord laminae at densities equivalent to those of GABAB1 in the dorsal horn. GABAB2 mRNA was also detected to the same degree within the DRG. Immunocytochemical analysis revealed that GABAB(1a), GABAB(1b) and GABAB2 were all present in the spinal cord. GABAB(1a) labelling appeared to be more dense than GABAB(1b) and within the superficial dorsal horn GABAB(1a) was present in the neuropil whereas GABAB(1b) was associated with cell bodies in this region. Both 1a and 1b immunoreactivity was expressed in motor neurons in lamina IX. GABAB2 immunoreactivity was expressed throughout the spinal cord and was evident within the neuropil of the superficial laminae.
Towers
S
S
Department of Pharmacology, Medical School, University of Birmingham, Birmingham, B15 2TT, UK.
Princivalle
A
A
Billinton
A
A
Edmunds
M
M
Bettler
B
B
Urban
L
L
Castro-Lopes
J
J
Bowery
N G
NG
D000818:Animals; D001418:Baclofen / Q000494:pharmacology; D001565:Benzoates / Q000494:pharmacology; D019281:Dimerization; D018755:GABA Agonists / Q000494:pharmacology; D005727:Ganglia, Spinal / Q000737:chemistry / Q000502:physiology*; D015870:Gene Expression / Q000502:physiology; D017403:In Situ Hybridization; D007536:Isomerism; D008297:Male; D009943:Organophosphorus Compounds / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis; D011869:Radioligand Assay; D051381:Rats; D018080:Receptors, GABA-B / Q000032:analysis / Q000737:chemistry / Q000235:genetics*; D017661:Receptors, Presynaptic / Q000032:analysis / Q000737:chemistry / Q000235:genetics; D013116:Spinal Cord / Q000737:chemistry / Q000502:physiology*; D014316:Tritium
D001565:Benzoates; C114540:CGP 62349; D018755:GABA Agonists; D009943:Organophosphorus Compounds; D012333:RNA, Messenger; D018080:Receptors, GABA-B; D017661:Receptors, Presynaptic; D014316:Tritium; D001418:Baclofen