Topographic organization of facial motoneurons to individual pinna muscles in rat (Rattus rattus) and bat (Rousettus aegyptiacus)

The location and number of motoneurons to individual pinna muscles were determined by retrograde transport of horseradish peroxidase in rat and flying fox. The degree of ear mobility differs considerably between these species in that rats perform simpler ear movements while flying foxes move their pinnae in a sophisticated way. Five pinna muscles were investigated in each species. Motoneurons lay within the medial subdivision of the facial motor nucleus extending over its entire rostrocaudal length. They were topographically organized; however, a somatotopic order could not be observed. With one exception homologous pinna muscles were represented in corresponding areas in both species, supporting the idea of a common representation of ear muscles in mammals. In rat, motoneuron pools overlapped considerably, whereas in flying fox overlap was minute. A total of 1,110 and 1,646 motoneurons were labeled in rat and flying fox, respectively. We conclude that the higher number of pinna motoneurons in the latter species in addition to the more clear-cut topography provide the structural substrates that underlie differences in the quality of ear movements as seen in bats vis-a-vis other mammals.     

The location of the preganglionic neurons that innervate the submandibular gland of the cat. A horseradish peroxidase study

Horseradish peroxidase was injected under pressure in the main excretory duct of the submandibular gland of cats, to locate the perikarya of the preganglionic parasympathetic salivatory neurons in the brainstem. Labeled multipolar neurons were found ipsilaterally in the lateral reticular formation, where they extended from the rostral plane that contains the genu of the facial nerve to the caudal plane that contains the rostral pole of the dorsal motor nucleus of the vagus nerve.     

Orexin B immunoreactive fibers and terminals innervate the sensory and motor neurons of jaw-elevator muscles in the rat

The relationship between orexinergic terminals and the sensory and motor neurons of jaw-elevator muscles was examined by means of anti-orexin B (OXB) immunohistochemistry combined with horseradish peroxidase (HRP) retrograde tracing in the rat. HRP was initially injected into the jaw-elevator muscles; 48 h later the animals were sacrificed and fixed for HRP reaction and anti-OXB immunohistochemistry. OXB-like terminals were observed to distribute in the trigeminal mesencephalic nucleus (Vme) and the trigeminal motor nucleus (Vmo) where they closely contact the Vme neuronal somata and the Vmo neuronal somata and dendrites retrogradely labeled with HRP. The results of this study provide anatomical evidence of a direct OXB orexinergic innervation of the sensory and motor neurons controlling jaw-elevator muscles involved in mastication. Its functional significance related to the feeding behavior and bruxism is discussed.     

Muscle units divided among retractor bulbi muscle slips and between the lateral rectus and retractor bulbi muscles in cat

Single muscle units of the retractor bulbi and lateral rectus muscles were activated with brief (0.5-ms) current pulses delivered through an intracellular micropipet penetrating single motoneuron cell bodies or axons in the principal abducens nucleus of the cat. Muscle unit mechanical characteristics were measured. A total of 40 retractor bulbi muscle units was studied. Thirty muscle units were contained in 2, 3, or all 4 retractor bulbi muscle slips. Three muscle units involved one retractor bulbi muscle slip and seven units were contained in the lateral rectus muscle plus one or both of the lateral retractor bulbi muscle slips. The largest percentage of muscle units (32.5%) was contained in the two inferior retractor bulbi muscle slips. The 33 retractor bulbi muscle units, having no muscle fibers in the lateral rectus muscle, caused the contraction of 76 retractor bulbi muscle slips. Twitch tension was measured in 58 slips and maximum tetanic tension in 27 slips. The average twitch tension per measured slip was 68.0 mg and the average maximum tetanic tension was 459.8 mg. The average retractor bulbi muscle unit contraction time was 10 ms and the lateral rectus average contraction time was 7.9 ms. We propose that peripheral branching of abducens nerve axons can account for the division of muscle unit components among the retractor bulbi muscle slips and between the lateral rectus and retractor bulbi muscles.     

Glutamate-like Immunoreactivity in Retinal Terminals of the Mouse Suprachiasmatic Nucleus

With a view to identifying the neurotransmitter content of retinal terminals within the mouse suprachiasmatic nucleus, a highly specific antiserum to glutaraldehyde-coupled glutamate was used in a postembedding immunogold procedure at the ultrastructural level. Retinal terminals were identified by cholera toxin–horseradish peroxidase transported anterogradely from the retina and reacted with tetramethyl benzidine/tungstate/H₂O₂, or by their characteristically pale and distended mitochondria with irregular cristae. Controls included model ultrathin sections containing high concentrations of various amino acids. Alternate serial sections were labelled with anti-glutamate and anti-γ-aminobutyric acid (GABA). Data were analysed by computer-assisted image analysis. Density of glutamate labelling (gold particles per μm²) on whole retinal terminals was > 3 times higher than that on postsynaptic dendrites, and > 5 times higher than that on miscellaneous non-retinal non-glutamatergic terminals in the suprachiasmatic nucleus. The overall density of gold particles over retinal terminals was ～ 3 times higher than that over GABAergic terminals, in which glutamate-like immunoreactivity was mainly mitochondrial. Labelling of vesicles in retinal terminals was almost 5 times greater than the apparent labelling of vesicles in GABAergic terminals, underscoring the location of transmitter glutamate within synaptic vesicles in retinal terminals. In the retino-recipient region of the suprachiasmatic nucleus there was also a small population of non-retinal glutamatergic terminals. Their overall immunoreactivity was similar to or exceeded that of retinal terminals, but morphological features clearly distinguished between these two types of glutamate-containing terminals. The present results indicate that the vast majority of retinal terminals may use glutamate as a transmitter, in keeping with electrophysiological and neuropharmacological data from other sources. The possibility of cotransmitters within retinal terminals, suggested by the presence of dense-core vesicles among the glutamate-containing synaptic vesicles, has still to be addressed.     

Morphology of rat spinal motoneurons with normal and hormonally altered specificity

Potential determinants of motoneuronal morphology were examined by using a sexually dimorphic, steroid-sensitive neuromuscular system in the rat spinal cord. In males, the spinal nucleus of the bulbocavernosus (SNB) innervates the perineal muscles bulbocavernosus (BC) and levator ani (LA), and the dorsolateral nucleus (DLN) innervates the ischiocavernosus muscle (IC). Adult females normally lack these motoneurons and the peripheral targets. Prenatal exposure of females to the androgen dihydrotestosterone propionate (DHTP) partially masculinizes this neuromuscular system and alters moto-neuron-to-muscle specificity, resulting in retained SNB target muscles anomalously innervated by motoneurons in the DLN. Because the morphology of SNB and DLN motoneurons normally differs significantly, the influence of spinal cord location and peripheral target on motoneuron morphology can be directly compared. Injection of cholera toxin conjugated to horseradish peroxidase (CTHRP) into the LA of DHTP-treated females labeled motoneurons predominantly in the SNB. These (SNB-LA) motoneurons in DHTP females were identical in all morphological measures to those of normal males. CTHRP injection into the BC of DHTP females labeled motoneurons in both the SNB and the DLN. SNB-BC motoneurons in DHTP females resembled those of normal males in process number and orientation, but were significantly smaller in dendritic length per motoneuron and in soma size. The DLN motoneurons anomalously projecting to the BC in DHTP females differed significantly from SNB-BC motoneurons in soma size and number and orientation of primary processes. However, these motoneurons were identical in all respects to DLN-IC motoneurons in DHTP females; DLN-IC motoneurons were similar to those of normal males in the orientation of their dendritic arbor, but were significantly smaller in dendritic length, soma size, and number of primary processes. These comparisons make it clear that DHTP selectively affects motoneuronal specificity and morphology in specific motoneuron classes. Further, motoneuronal morphology in the SNB/DLN system appears to be influenced more by spinal cord location than by peripheral target.     

Morphology of single primary spindle afferents of the intercostal muscles in the cat

A reconstruction was made of the trajectory of primary spindle afferents from the intercostal muscles in the spinal cord of the cat. Intraaxonal recordings were performed from the primary spindle afferents that were identified by their response to lung inflation and stimulus threshold to activate the action potentials. The afferents were stained by using intraaxonal injection of horseradish peroxidase (HRP). Results were obtained mainly from internal intercostal Ia fibers, which entered the spinal cord and bifurcated into ascending and descending branches. The ascending branches could be traced up to 10.7 mm, and the descending branches could be traced up to 7.3 mm. The ascending branches extended to the next segment. Collaterals ranging from one to six were given off from these branches. The distances between adjacent collaterals ranged from 0.9 mm to 4.7 mm. Each collateral had similar morphological characteristics. The collaterals entered the dorsal horn and ran toward lamina IX through the medial half of the gray matter. Fine branches and boutons were given off in laminae V, VII, VIII, and IX. The aggregations of these branches were found in lamina VII, mainly in the region of Clarke's column and in the ventral and ventrolateral regions thereof and in lamina IX, mainly in the nucleus lateromedialis. Most terminals did not contact the somata of target neurons in all laminae in which terminals were found. However, a few terminals were found to contact large neurons in lamina IX. In addition to these aggregates, there were some terminals scattered throughout the ventral horn. Thus, it was concluded that single intercostal Ia afferents project to the region of Clarke's column, to the intercostal motor nucleus, and to the intermediate regions. J. Comp. Neurol. 398:459–472, 1998. © 1998 Wiley-Liss, Inc.     

Organizational and morphological features of cat sacrocaudal motoneurons.

We have investigated the organizational and morphological features of motoneurons from cat sacrocaudal spinal cord, the portion of the neuraxis that innervates the tail. This information is pertinent for development of a new model of spinal cord injury. An understanding of sacrocaudal circuitry is essential for physiological and behavioral assessment of the effects of sacrocaudal lesions. Observations from Nissl-stained sections corroborated Rexed's cytoarchitectural scheme. Putative motoneurons were located within two regions of the ventral horn: the ventromedial nucleus (lamina IX) and the nucleus commissuralis. To map motoneuron pools, cholera toxin-horseradish peroxidase conjugate was injected into each dorsal tail muscle. The dorsomedial muscle was innervated by ipsilateral nucleus commissuralis motoneurons. The dorsolateral and intertransversarius muscles were innervated by ipsilateral lamina IX and nucleus commissuralis motoneurons. Cell bodies of retrogradely labeled sacrocaudal motoneurons ranged from 22 to 82 microns in diameter; the unimodal distributions peaked between 45 and 50 microns. Dendritic trees of motoneurons, revealed by retrograde labeling or by intracellular injection with horseradish peroxidase, were extensive. Five to eight primary dendrites originated from the cell body. Dendritic branches extended throughout the ipsilateral ventral gray matter, with processes spreading into the surrounding white matter and the base of the dorsal horn. Dendrites from motoneurons with their soma in the lateral portion of lamina IX formed a longitudinal plexus at the gray/white border. Medial dendrites from motoneurons in the nucleus commissuralis formed bundles in the ventral gray commissure and spread throughout the contralateral ventral horn. It is speculated that contralateral dendrites subserve synchronized co-contraction of medial muscles from both sides of the tail.     

An HRP study of the central projections of primary trigeminal neurons which innervate tooth pulps in the cat

Trigeminal ganglia and brain stem of adult cats were studied following HRP injections into tooth pulps or after exposure of the cut end of the inferior alveolar nerve to HRP. Ipsilateral ganglion cells within a wide range of sizes were labeled in both experimental situations, whereas no labeled cells were observed in the contralateral ganglion in any animal. Labeled central branches of tooth pulp and inferior alveolar neurons were observed in all subdivisions of the ipsilateral trigeminal sensory complex. Terminal labeling in the tooth pulp experiments was confined to the dorsomedial parts of the main sensory nucleus and subnuclei oralis and interpolaris. Caudal to the obex terminal labeling was restricted to the medial halves of laminae I, IIa and V of the medullary dorsal horn. In the inferior alveolar nerve experiments dense terminal labeling was observed in the dorsal parts of the main sensory nucleus and subnuclei oralis and interpolaris. Caudal to the obex terminal labeling was located throughout laminae I to V in contrast to the tooth pulp experiments. Neither of the two experimental situations offers any evidence for a bilateral or contralateral brain stem projection of primary trigeminal neurons.     

Patterns of primary afferent termination in the external cuneate nucleus from cervical axial muscles in the cat

Using the method of transganglionic transport of horseradish peroxidase (HRP), the distribution of primary afferent projections was examined in the external cuneate nucleus (ECN) from different muscle groups in the forequarter of the cat. The terminal zones of afferent fibers from three shoulder muscles--clavotrapezius, acromiotrapezius, and spinotrapezius--were compared to projections from suboccipital muscles, dorsal neck extensors, and muscles of the proximal forelimb. Each muscle group had a labelled terminal zone that occupied a different subvolume of the ECN. The zone labelled from trapezius muscles formed a continuous column in the ECN running from the caudal pole of the nucleus to a level 3.0 mm rostral to the obex. Terminal zones of suboccipital muscles and dorsal neck extensors formed longer columns that extended into the most rostral tip of the ECN, while those of proximal forelimb muscles formed shorter columns confined to the caudal two-thirds of the ECN. At comparable cross-sectional levels in the caudal and middle portions of the ECN, terminal zones from proximal limb muscles were located most dorsomedially, while those from shoulder muscles, dorsal neck muscles, and suboccipital muscles were located in progressively more ventral and lateral regions. The subvolume of the ECN occupied by projections from cervical axial muscles was estimated to be more than 40% of the volume of the nucleus, suggesting that the ECN has a major role in the transmission of sensory information from axial musculature to the cerebellum. Following exposure of all muscle nerves to tracer, a second labelled zone was also identified close to the ECN in the descending vestibular nucleus at transverse levels 2.0-3.0 mm rostral to the obex. Here, reaction product was concentrated around a circumscribed collection of medium-sized, multipolar cells.     

Distribution of motoneurons supplying dorsal suboccipital and intervertebral muscles in the cat neck

Multiple labelling methods were used to examine the topographical organization of motoneurons supplying the suboccipital muscles, rectus capitis posterior (RCP) and obliquus capitis inferior (OCI); the intervertebral muscles, spinalis dorsi (SD) and semispinalis cervicis (SSC); and the dorsal extensor biventer cervicis (BC). Muscle nerves were isolated, cut, and exposed in the same animal to one of three different retrograde tracers, Fluorogold, Fast Blue, and horseradish peroxidase. Motoneurons supplying suboccipital muscles were found to be located in the rostral two cervical segments. The RCP motor nucleus was confined to C1 and the caudal brainstem, whereas the OCI nucleus usually originated in mid-C1 and extended caudally as far as mid-C2. Most motoneurons supplying OCI and SD were concentrated in the deepest part of the ventral horn, but some cells were found in the commissural region, the contralateral ventral horn, the dorso-medial aspect of the ipsilateral ventral horn, and the ventromedial white matter. Cells supplying the functional extensor RCP were intermingled with those supplying the turning muscle OCI, although a gradient in mediolateral distribution was detected. Motoneurons supplying the intervertebral muscles SD and SSC were found caudal to C3. Within the ventral horn, they occupied locations similar to those occupied by suboccipital motoneurons in C1 and C2. The cell column for intervertebral motoneurons was just medial to that of BC and some cells supplying SD and SSC were found in the territory occupied by BC motoneurons. Motoneurons supplying intervertebral and suboccipital muscles had unimodal diameter spectra, and most cells had mean equivalent diameters smaller than 35 microns.     

Auditory brainstem projections to the ferret superior colliculus: Anatomical contribution to the neural coding of sound azimuth

The mammalian superior colliculus (SC) contains a neural map of auditory space. It is not known whether this topographic representation emerges at the level of the SC or is relayed there from other auditory areas. We have used retrograde labelling techniques in ferrets to examine the sources and pattern of innervation from auditory brainstem nuclei. After multiple injections of wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP) into the SC, the heaviest concentrations of labelled cells were found in the nucleus of the brachium (BIN) and external nucleus of the inferior colliculus, with much weaker labelling in the nucleus sagulum, dorsal, intermediate and ventral nuclei of the lateral lemniscus, paralemniscal regions, and periolivary nuclei. The projections were predominantly ipsilateral, although labelled cells were found on both sides of the brainstem. Single injections of WGA-HRP or discrete injections of red and green latex microspheres revealed that the caudal and lateral regions of the SC receive the heaviest projections, although the majority of the retrogradely labelled neurons in the contralateral BIN project to rostral SC. On the ipsilateral side, neurons in rostral and caudal regions of the BIN were labelled primarily by the tracer injected into rostral and caudal regions of the SC, respectively. However, no clear segregation was apparent in the BIN after injections into the medial and lateral regions or in any of the other nuclei after either injection paradigm. These data suggest that converging inputs from several auditory brainstem nuclei contribute to the construction of the auditory space map in the SC, although information about sound azimuth may be conveyed to this nucleus via a spatially ordered projection from the ipsilateral BIN. J. Comp. Neurol. 390:342–365, 1998. © 1998 Wiley-Liss, Inc.     

Trigeminal nerve and temporomandibular joint of the cat: A horseradish peroxidase study

The mesencephalic nucleus of the trigeminal nerve is considered to contain the cell bodies of the first-order neurons that have peripheral connections in the temporomandibular joint capsule. Through use of retrograde transport of horseradish peroxidase, this concept is challenged. The results indicate that a relatively specific region of the mandibular division of the trigeminal ganglion of the cat contains the first-order neurons innervating the temporomandibular joint capsule.     

A qualitative investigation of the topographical representation of masticatory muscles within the motor trigeminal nucleus of the rat: a horseradish peroxidase study

The topographical representation of the masticatory muscles of the rat was investigated by studying retrograde transport of horseradish peroxidase (HRP) from individual muscles. Contrary to the classically accepted scheme, the temporalis and masseter are separately represented dorsolaterally, the pterygoids dorsomedially and the jaw-opening mylohyoid and anterior belly of digastric ventrally within the motor nucleus, corresponding to the arrangement of the muscles on the head. Phylogenetic and ontogenetic justifications for this organization are adduced.     

Distribution and morphology of sacral autonomic neurons in the cat: peroxidase labeling of efferent neurons through transected ventral roots.

The distribution and morphologic characteristics of the nucleus intermediolateralis inferior, the cell group X of Onuf, and the medial border cell group of the ventral horn, the latter two of which are also possibly autonomic in function, were investigated in the cat by means of retrograde axonal transport of horseradish peroxidase through the cut end of the transected individual sacral ventral root. The nucleus intermediolateralis inferior contributes its axons chiefly to the S2 and S3 ventral roots, and the nucleus occupies a larger triangular area of the intermediate region than that described previously. The neurons of the intermediolateral group are arranged in horizontal fashion. Cell group X contributes its axons chiefly to S1, and this is composed of longitudinally arranged, medium-size neurons having very well-developed longitudinal dendritic bundles. The medial border cells of the ventral horn, which are chiefly of medium and small size, are also labeled by bathing the S1 through C×1 roots. Their neuronal characteristics suggest that they might be also autonomic in function.     

Ultrastructural analysis of enkephalinergic terminals in parasympathetic ganglia innervating the urinary bladder of the cat

Leucine enkephalin immunoreactivity was identified in axons and varicosities in parasympathetic ganglia located in the pelvic plexus and on the surface of the urinary bladder of the cat. Electron microscopic immunohistochemical studies revealed that varicosities containing leucine enkephalin exhibited large dense core vesicles and small, clear, spherical vesicles, which were similar to those found in cholinergic terminals. Leucine enkephalin immunoreactivity was primarily associated with large dense core vesicles. The varicosities formed axodendritic and axosomatic synapses with principal ganglion cells. Axoaxonic synapses were not detected. Some axosomatic enkephalinergic synapses were detected embedded within or invaginating the principal ganglion cells. Varicosities containing flattened and/or small dense core vesicles did not exhibit enkephalin immunoreactivity. Bladder ganglion cells identified by retrograde HRP tracing from the urinary bladder exhibited similar leucine enkephalinergic synapses. These observations, coupled with previous reports that leucine enkephalin is present in sacral preganglionic neurons and released by preganglionic nerve stimulation, suggest that leucine enkephalin and acetylcholine are cotransmitters stored and released from the same nerve terminals in bladder parasympathetic ganglia.     

Is there morphological separation between the spinal cord motor nuclei which innervate the heads of a multiheaded muscle?

Horseradish peroxidase was injected into the individual heads of the pectoralis muscles of the dog or applied to the nerve which supplies each of these heads. The location and numbers of labeled motoneurons in the spinal cord were studied using light microscopy. There was longitudinal overlap of the pectoral nuclei, but no separation in their mediolateral or dorsoventral positions. The cutaneous trunci muscle motor nucleus is distinctly separate from the motor nuclei of the pectoral muscles, even though they share a common nerve supply. The methods of horseradish peroxidase application to the cut nerve or injection into the muscle are compared.     

Structural changes of the soleus and the tibialis anterior motoneuron pool during development in the rat.

The morphological development of motoneuron pools of two hindlimb muscles of the rat, soleus (SOL) and tibialis anterior (TA), was studied in rats ranging in age between 8 and 30 postnatal days (P8-P30). Motoneurons were retrogradely labelled by injecting a cholera toxin B subunit solution directly into the muscles. This resulted in extensive labelling of motoneurons as well as their dendritic trees. The distribution of cross sectional areas of neuronal somata was determined for both muscles at various ages. Somal size increased considerably between P8 and P12, whereas growth was moderate between P12 and P20. The size distribution of SOL motoneurons was bimodal from P20, whereas the size distribution of TA motoneurons remained largely unimodal. The morphological development of the dendritic tree was studied qualitatively. The development of dendritic arborization within the SOL and the TA motoneuron pool showed major differences. The arborization pattern of dendrites of TA motoneurons was basically multipolar at all ages. In contrast, dendrites of SOL neurons tended to line up with the rostro-caudal axis and became organized in longitudinal bundles from P16 onwards. The relatively late appearance of dendrite bundles in the soleus motoneuron pool suggests that they might be related to the fine-tuning of neuronal activity rather than patterning of motor activity. The occurrence of dendrite bundles in SOL and not in TA motoneuron pools suggests that they may be related to the different afferent organization of this postural muscle or to its tonic activation pattern.