Distribution of immunoreactivity for the β ₂ and β ₃ subunits of the GABA _A receptor in the mammalian spinal cord

Abstract

The localization of GABAA receptors in cat and rat spinal cord was analyzed using two monoclonal antibodies specific for an epitope shared by the beta 2 and beta 3 subunits of the receptor. beta 2/beta 3-subunit immunoreactivity was the most intense in inner lamina II, lamina III, and lamina X, and it was the least intense in lamina IX. In laminae I-III, generally, the staining had a rather diffuse appearance, but the surfaces of small cell bodies in these laminae were outlined clearly by discrete labeling, as were many cell bodies and dendrites in deeper laminae. Rhizotomy experiments and ultrastructural observations indicated that beta 2/beta 3-subunit immunoreactivity in the dorsal horn was largely localized in intrinsic neuropil elements rather than in the terminals of primary afferent fibers, even though labeling overlapped with the terminal fields of different types of primary afferents and was also detected on the membranes of dorsal root ganglion neurons. With few exceptions (most notably, a highly immunoreactive group of dorsolaterally located cells in the cat lumbar ventral horn), motoneurons expressed low levels of beta 2/beta 3-subunit immunoreactivity. Labeling of neuronal membranes was fairly continuous, but focal accumulations of beta 2/beta 3-subunit immunoreactivity were also detected using immunofluorescence. Focal "hot spots" correlated ultrastructurally with the presence of synaptic junctions. Dual-color immunofluorescence revealed that focal accumulations of beta 2/beta 3-subunit immunoreactivity were frequently apposed by glutamic acid decarboxylase (GAD)-immunoreactive terminals. However, the density of continuous-membrane beta 2/beta 3 immunolabeling and GAD terminal density were not correlated in many individual neurons. The results suggest the existence of "classical" (synaptic) and "nonclassical" (paracrine) actions mediated via spinal cord GABAA receptors. The study also revealed the relative paucity of beta 2/beta 3-subunit immunoreactivity postsynaptic to certain GABAergic terminals, particularly those presynaptic to motoneurons or primary afferent terminals.

GABA receptors modulate trigeminovascular nociceptive neurotransmission in the trigeminocervical complex

“…In situ studies indicate that intrinsic neurones contribute signi®cantly to the GABA A receptor population in the dorsal horn. Ultrastructural studies show that GABA A receptors usually participate in classical synapses and GABA A -immunoreactivity has been found at post-synaptic specializations in the spinal cord (Alvarez et al, 1996;Coggeshall & Carlton, 1997). L-Glutamate is major source of excitatory transmission in the CNS (Seeburg, 1993).…”

A spinal GABAergic mechanism is necessary for bladder inhibition by pudendal afferent stimulation

“…GABA A receptors are located throughout the dorsal horn and in the dorsal gray commissure (DGC) (2). GABA A receptors also occur on neurons in the SPN (3), whose dendrites extend to the DGC (13,30), a site of pudendal afferent innervation (49).…”

“…GABA A receptors also occur on neurons in the SPN (3), whose dendrites extend to the DGC (13,30), a site of pudendal afferent innervation (49). There may be multiple inhibitory pathways to the SPN (17), including both presynaptic and postsynaptic GABAergic inhibition from other areas of the sacral spinal cord (2,35). Inhibitory feedback neurons from the SPN are known to produce recurrent inhibition of the bladder circuit (12) and may be GABAergic.…”

Differential Plasticity of the GABAergic and Glycinergic Synaptic Transmission to Rat Lumbar Motoneurons after Spinal Cord Injury

“…Immunocytochemistry showed a complete absence of immunolabeling of the GABA A R ␣1, ␣2, ␣3, and ␣5 subunit, in each respective mice (Kralic et al, 2006). The monoclonal mouse antibody bd-17 (US Biological, bovine, cat # G1016; 1:400) directed against both ␤2 and ␤3 subunits of GABA A Rs, recognize the major GABA A R subtypes in the CNS (McKernan and Whiting, 1996) expressed during postnatal development (Alvarez et al, 1996) and in adult spinal cord motoneurons (Fritschy et al, 1994;Bohlhalter et al, 1996). The mature form of the GlyR, GlyR␣1, was detected with either the monoclonal antibody mAb2b (Connex, mouse, 1:80, catalog # 28 120 501-0 and Synaptic Systems (SySy), mouse 1:250, catalog # 146 111) or the polyclonal antibody pAb2b (Millipore Bioscience Research Reagents, rabbit, 1:100; catalog # AB5052), depending on the other primary antibodies used for double immunofluorescence staining (Pfeiffer et al, 1984;Schröder et al, 1991;Liu and Wong-Riley, 2002;Baer et al, 2003).…”

“…They were compared with three SCT-P7 rats which received equivalent volumes of NaCl 0.9% and with control P7 rats. To get an overall view of the effects of DOI on the GABA A R, bd-17 antibody, which is directed against both ␤ 2 and ␤ 3 subunits and therefore recognize the major GABA A R subtypes in the CNS (Fritschy et al, 1994;Alvarez et al, 1996;McKernan and Whiting, 1996) was use and covisualized with the GlyR␣1 on FBlabeled motoneurons (Fig. 6).…”