Confocal immunofluorescence microscopy showed strong monocarboxylate transporter 2 (MCT2) labeling of Purkinje cell bodies and punctate labeling in the molecular layer. By immunogold cytochemistry, it could be demonstrated that the MCT2 immunosignal was concentrated at postsynaptic densities of parallel fiber-Purkinje cell synapses. The distribution of MCT2 transporters within the individual postsynaptic densities mimicked that of the delta2 glutamate receptor, as shown by use of two different gold-particle sizes. The MCT2 distribution was also compared with the distributions of other monocarboxylate transporters (MCT1 and MCT4). The MCT1 immunolabeling was localized in the endothelial cells, while MCT4 immunogold particles were associated with glial profiles, including those abutting the synaptic cleft of the parallel fiber-spine synapses. The postsynaptic density (PSD) molecules identified so far can be divided into five classes: receptors, their anchoring molecules, molecules involved in signal transduction, ion channels, and attachment proteins. Here, we provide evidence that this list of molecules must now be extended to comprise an organic molecule transporter: the monocarboxylate transporter MCT2. The present data suggest that MCT2 has specific transport functions related to the synaptic cleft and that this transporter may allow an influx of lactate derived from perisynaptic glial processes. The expression of MCT2 in synaptic membranes may allow energy supply to be tuned to the excitatory drive.

Motor nerve terminals and adjacent structures in the extensor digitorum longus and soleus muscles of young adult rats were examined for their content of glutamate by means of quantitative, electron microscopic immunocytochemistry employing colloidal gold particles as markers. The level of glutamate immunoreactivity was stronger in the extensor digitorum longus terminals than in the soleus terminals. In both muscles the glutamate immunolabelling was stronger in the nerve terminals than in the synaptic clefts and the postsynaptic tissue separating the secondary clefts, but the differences were larger in the extensor digitorum longus than in the soleus muscle. The myofibrils of the soleus muscle were more densely labelled than those in the extensor digitorum longus muscle. The level of immunoreactivity was high in the Schwann cells of both muscles. By comparing the labelling intensity of motor nerve terminals with that of muscle fibres and hippocampal mossy fibres (compartments that have been analysed previously with respect to their glutamate content), the mean concentration of fixed glutamate in the extensor digitorum terminals was estimated to be in the range of 10-20 mmol/l. An association of glutamate immunoreactivity with synaptic vesicles was demonstrated in the most strongly labelled terminals. Whether these epitopes were localized in the interior of the vesicles or at their external surface could not be resolved with the present technique. These data indicate that motor nerve terminals contain glutamate, and that the enrichment of this amino acid is more pronounced in the terminals of the extensor digitorum longus muscle (a fast muscle) than in those of the soleus muscle (a slow muscle). A possible modulatory or trophic role of glutamate in the mammalian neuromuscular junction should be considered.

Neuromuscular junctions on fast and slow skeletal muscle fibers have different properties. Possible reasons for these differences were examined in adult rat soleus (SOL) muscle fibers reinnervated at new ectopic or old denervated sites by fast fibular (FIB) or slow SOL motoneurons. FIB motoneurons formed large ectopic junctions with a high density of nerve terminal varicosities (fast appearance), whereas SOL motoneurons formed small ectopic junctions with a low density of varicosities (slow appearance). Both FIB and SOL motoneurons formed small junctions with a slow appearance when reinnervating old SOL endplates. FIB nerves innervating ectopic sites and SOL nerves reinnervating old sites had the same appearance whether they contacted the SOL fibers alone (single innervation) or together (dual innervation). Continuous stimulation of the FIB nerve at 10 Hz for 3-4 months reduced the size of ectopic FIB and intact extensor digitorum longus (EDL) junctions and caused a modest reduction in density of terminal varicosities in EDL. Junction size and muscle fiber diameter were positively correlated, but the slope describing this relation was steeper for FIB junctions than for SOL junctions. It is concluded that in the present system (1) motoneuron type and not muscle fiber type determines the fast or slow character of the neuromuscular junction. (2) denervated endplates of one type place stable and severe constraints on the termination pattern of reinnervating axons of another type, (3) the appearance of fast EDL junctions undergoes a modest fast to slow transformation when exposed to long-term slow pattern stimulation, and (4) not only the size of the muscle fibers, but also the type and firing pattern of the motoneurons and the spatial constraints at preformed endplates influence the relation between junction size and muscle fiber diameter.