Quantitative analysis of synaptic contacts made between functionally identified oralis neurons and trigeminal motoneurons in cats

Abstract

A previous study revealed that rostrodorsomedial oralis (Vo.r) neurons synapsing on trigeminal motoneurons use GABA and/or glycine as neurotransmitters. To determine the number and spatial distribution of contacts, injections of biotinamide and horseradish peroxidase were made into a Vo.r neuron and an alpha-motoneuron in the jaw-closing (JC) and jaw-opening (JO) motor nucleus, respectively, in 39 cats. All Vo.r neurons responded to low-threshold mechanical stimulation of the oral tissues. Single Vo.r neurons terminating in the JC nucleus (Vo.r-dl neurons; n = 5) issued, on average, 10 times more boutons than Vo.r neurons terminating in the JO nucleus (Vo.r-vm neurons; n = 5; 4437 vs 445). The Vo.r-dl neuron-JC alpha-motoneuron pairs (n = 4) made contacts on either the soma-dendritic compartment or dendrites, and the Vo.r-vm neuron-JO motoneuron pairs (n = 2) made contacts on dendrites, with a range of two to seven contacts. In five of the six pairs, individual or groups of two to three terminals contacted different dendritic branches of a postsynaptic cell. The Vo.r-dl neurons innervated a greater number of counter-stained motoneuronal somata than did the Vo.r-vm neurons (216 vs 26). Total number of contacts per Vo.r neuron was higher for the Vo.r-dl than Vo.r-vm neurons (786 vs 72). The present study demonstrates that axonal branches of Vo.r neurons are divided into two types with different innervation domains on the postsynaptic neuron and that they are highly divergent. The overall effect exerted by these neurons is predicted to be much greater within the JC than JO motoneuron pool.

Fig. 1.

Physiology and morphology of a labeled neuron CL3 in the rostrodorsomedial part (Vo.r) of oral nucleus (A–C) and the somal location of labeled Vo.r neurons examined (D, E).A, Eight superimposed traces of intracellular potentials recorded from the Vo.r neuron CL3 after stimulation of the infraorbital nerve with submaximal intensity.B, Camera lucida drawings of soma-dendrites and part of the stem axon (arrowheads) of the Vo.r neuron.C, Diagram of axonal trajectory of the Vo.r neuron with terminals in the dorsolateral division (Vmo.dl) of the trigeminal motor nucleus (Vmo) and in the brainstem nuclei other than the Vmo.dl. Contacts made between the Vo.r neuron and labeled masseter motoneuron are marked with arrowheads labeled by S1, S2, and S3. The collaterals from an ascending and a descending axon are denoted bya and d, respectively. Xindicates the midline. D, E, Somal location of Vo.r neurons examined is plotted in two sections at the rostral (D) and caudal (E) levels of the Vo.r. Circles and squaresrepresent Vo.r neurons that project to the Vmo.dl and to the ventromedial division (Vmo.vm) of the Vmo, respectively.Arabic numerals within the circles andsquares indicate sample (neuron) numbers.SO, Superior olive; Vint, intertrigeminal region; Vjux, juxtatrigeminal region; Vp, trigeminal principal nucleus; Vpv, ventrolateral subnucleus of Vp; Vtr, spinal trigeminal tract;7N, facial nerve; D–M, dorsal–medial. Calibration: A, 10 mV, 4 msec. Scale bars: B, 2 mm; D, E, 1 mm.

Fig. 2.

Diagrammatic summary showing the locations of contacts made by four Vo.r-dl neuron–JC motoneuron pairs (A–D) and two Vo.r-vm neuron–JO motoneuron pairs (E, F), with terminal boutons denoted by open triangles and en passant boutons denoted by open circles. JC and JO motoneurons receiving contacts from single Vo.r neurons are marked with CL andOP, respectively. In each pair, contacts marked withSn are arbitrary. Scale bar, 100 μm (refers to the geometric dendritic distance from the soma). The distance was measured from reconstructions in the transverse plane and corrected by using the Pythagorean theorem and the section thickness. The a,d, and u indicate collaterals given off from an ascending fiber, a descending fiber, and a stem axon, respectively. Dendrites longer than 700 μm inA–C, E, andF and those longer than 1000 μm in Dare interrupted with a broken line. Arabic numeralsattached to the end of dendritic lines indicate the longest distance (μm) of the dendritic tree formed by each primary dendrite. Note that the second dendritic line inD starts 550 μm from the soma. The axon collateral branching patterns and somata are also shown, but not to scale. For further descriptions, see Results.

Fig. 3.

Camera lucida drawings reconstructed from a labeled masseter motoneuron (blue) and a labeled Vo.r neuron CL3 (red) showing three contacts made between the two. Contacts are marked with arrows labeled with S1, S2, and S3. The size of labeled boutons drawn is exaggerated. Note that the intracellular responses, soma-dendrites and a stem axon, and scheme of the axonal trajectory of the Vo.r neuron are shown in Figure1A–C, respectively.D–M, Dorsal–medial. Scale bar, 0.4 mm.

Fig. 4.

Parts of drawings reconstructed from a labeled Vo.r neuron CL4 and a labeled masseter motoneuron showing six contacts (S1–5 and S7) and photomicrographs of the contacts S1 and S7. A, Contacts made by labeled boutons from the Vo.r neuron on the soma (S1, S2, and S3,arrows) and a primary dendrite (S4 andS5, filled arrowheads) of the labeled masseter motoneuron and on a counter-stained motoneuronal soma (open arrowheads) in the Vmo.dl. Photomicrograph in theinset shows the contact S1(arrow). B, Contacts made by a labeled bouton on a proximal dendrite (S7, filled arrowhead) of the labeled motoneuron and two labeled boutons on the soma of a counter-stained motoneuron (open arrowheads) in the Vmo.dl. Photomicrograph in theinset shows the contact S7(filled arrowhead). A contact S6 is not included in sections used for the reconstructions. Note that the labeled soma and the counter-stained somata are cut into two pieces, and each contact is seen on the surface of a smaller piece. D–M, Dorsal–medial. Scale bars: reconstructions A andB, 20 μm; insets A and B, 5 μm.

Fig. 5.

Camera lucida drawings reconstructed from a labeled jaw-opening (JO) motoneuron and a labeled Vo.r neuron OP1 showing six contacts made between the two. Contacts are marked witharrows labeled with S1,S2, S3, S4,S5, and S6. The size of labeled boutons drawn is exaggerated. D–M, Dorsal–medial. Scale bar, 0.4 mm.

Fig. 6.

Parts of drawings illustrated in Figure 5 showing the contacts S3 and S4(A) and the contacts S5 andS6 (B) at higher magnification and photomicrographs of the contacts S3 and S6. A, Contacts made by two labeled boutons on the same dendrite of the labeled JO motoneuron (S3 and S4,filled arrowheads). Photomicrograph in theinset shows the contact S3(filled arrowhead). B, Contacts made by two labeled boutons on the same dendrite of the labeled motoneuron (S5 and S6, filled arrowheads). Photomicrograph in the inset shows the S6 contact (filled arrowhead). Note that labeled boutons make contacts with the soma of two counter-stained JO motoneurons (open arrowheads). D–M, Dorsal–medial. Scale bars: reconstructions A and B, 20 μm; insetsA and B, 5 μm.

Fig. 7.

Distribution patterns of labeled boutons from a Vo.r neuron CL1 in the Vmo.dl (Vo.r-dl neuron; A–C) and from a Vo.r neuron OP1 in the Vmo.vm (Vo.r-vm neuron; D–F). The Vo.r-dl neuron boutons found in every alternate section at levels of the rostral (A), middle (B), and caudal one-third (C) of the Vmo are superimposed in one representative section, respectively. The Vo.r-vm neuron boutons found in serial sections at the rostral (D), middle (E), and caudal (F) levels of the Vmo are superimposed in one representative section, respectively. Boutons contacting the somata and/or juxtasomatic regions of counter-stained motoneurons are marked with large dots. Boutons without contacts on the counter-stained motoneurons are marked with small dots. Dor–Med, Dorsal–medial. Scale bar, 0.5 mm.

Fig. 8.

Photomicrographs showing contacts made by a labeled Vo.r-dl neuron CL1 on the soma (A) and the juxtasomatic region (B) of counter-stained motoneurons. Filled arrowheads denote contacts that are relatively in focus. Contacts marked with open arrowheads are out of focus but could be identified by adjusting the focus. Scale bar, 20 μm.

Fig. 9.

Size distributions of counter-stained somata without contacts (open columns) and with contacts (filled columns) in the JC (A) and the JO (B) motor nucleus. The somata measured were randomly selected from sections in case Vo.r-dl neuron CL5 and in case Vo.r-vm neuron OP3.