Distribution of glutamate-, glycine- and GABA-immunoreactive nerve terminals on dendrites in the cat spinal motor nucleus

Abstract

The dendritic tree constitutes more than 93% of the receptive membrane area of a spinal motoneuron, yet little is known about its synaptic inputs. In this study we examined the distribution of glutamate-, GABA- and glycine-like immunoreactivity in boutons apposing dendrites in the L7 spinal cord motor nucleus, by use of postembedding immunohistochemistry on serial sections. We examined 799 boutons apposing 401 cross-sectioned dendrites of different calibre (range 0.2-15 microm), and 14 first-order (stem) dendrites. Thirty-five percent (35%) of the boutons were immunopositive for glutamate and 59% for GABA and/or glycine. Among the latter, 30% showed glycine immunoreactivity only and 24% were immunoreactive for both GABA and glycine. Very few were immunoreactive only for GABA (5%). As few as 6% of the boutons were judged as not enriched for any amino acid analysed. The fine structural characteristics of the boutons were in accordance with previous descriptions. The sample of dendrites was arranged in calibre bins in order to facilitate distribution analysis. Stem dendrites differed from the other bins, with a high total bouton covering (61%) and a high bouton density. Sixty-nine percent of the membrane covering was by glycine- and/or GABA-immunoreactive boutons, whereas 18% was covered by boutons enriched in glutamate. For non-stem dendrites, bouton covering fell from 33% to 12% with decreasing calibre. However, bouton apposition length decreased in parallel, yielding a fairly uniform bouton density among dendrites of different calibre. The lack of correlation between packing density and dendrite calibre was also evident when the sample of dendrites was broken down into subsamples based on content of amino acid immunoreactivity. The latter analysis also revealed that both the relative covering and density of boutons containing inhibitory amino acids (57%; glycine and/or GABA) and glutamate (38%), respectively, did not vary systematically with dendrite calibre. Combined, the data indicate that in non-stem dendrites the proportion of excitatory and inhibition inputs does not change systematically throughout the dendritic arborizations of spinal alpha-motoneurons. Thus, spinal motoneurons can, with respect to the general synaptic architecture, be divided into two main compartments, i.e. the proximal soma-juxtasomatic compartment (including stem dendrites) and the distal dendritic compartment. The proximal domain is under a powerful glycine and/or GABA influence. Finally, based on the data presented here and previously published data, it was calculated that spinal alpha-motoneurons receive in the range of 50-140 x 10(3) synaptic boutons.

Synaptic Control of Motoneuronal Excitability

“…The dendritic membrane constitutes >97% of the total membrane surface area in cat spinal motoneurons (246), with 61% of the stem dendrites and 12-33% of more distal dendrites covered by synaptic boutons (937). Consequently, the vast majority of synaptic inputs to motoneurons are dendritic, and integration of synaptic potentials is heavily influenced by the passive membrane properties of the dendrites (548,1024).…”

“…About 6% of the dendritic boutons are not immunoreactive for GABA, glycine, or glutamate. Presumably these boutons contain other transmitters (937).…”

“…Data are most consistent with Lglutamate as the main EAA at motoneuron synapses. Circumstantial evidence includes presence of high-affinity uptake mechanisms and high glutamine levels (456,1151) in neurons and terminals that synapse on motoneurons (144,808); detection of increased glutamate levels in perfusate from in vitro preparations following synaptic activation or glutamate uptake inhibition in rat and frog spinal cord (443,614,1223); potentiation of endogenous activity by EAA uptake inhibitors (142,442,818); and immunohistochemical detection of glutamate in boutons synapsing on motoneurons (878,937,1207 The functional complexity of glutamate receptor-mediated synaptic signaling is conferred by variation in receptor subunit composition and further enhanced by posttranscriptional processing of gene transcripts through alternative splicing and RNA editing. All AMPA receptor subunits undergo alternative splicing of COOH-terminal sequences and flip and flop sequences near the M4 transmembrane domain.…”

GABA‐, glycine‐, and glutamate‐immunoreactive bouton profiles in apposition to neurons of the central cervical nucleus in the rat

“…In accordance with results of previous studies (e.g., Ö rnung et al., 1996, 1998Lindå et al, 2000), glycine-ir bouton profiles contained flat vesicles and formed symmetric synaptic complexes ( Fig. 3A and B), whereas GABA-ir profiles contained either flat (Fig.…”

“…The corresponding ratio of gold particle density for glutamate was 3 or more (Ö rnung et al, 1998(Ö rnung et al, , Lindå et al, 2000. The rationale for using these ratios is that in bouton profiles with a labeling ratio in excess of 3 (glutamate) or 5 (GABA and glycine), a positive correlation is seen between gold particle density and number of vesicles (see also Fig.…”