J.C.V.M. Copray

Projections from the rostral parvocellular reticular formation to pontine and medullary nuclei in the rat: Involvement in autonomic regulation and orofacial motor control

The efferent connections of the rostral parvocellular reticular formation to pontine and medullary nuclei in the rat were studied with anterogradely transported Phaseolus vulgaris leucoagglutinin. Dense innervations from the rostral parvocellular reticular formation were found in the mesencephalic trigeminal nucleus, the supratrigeminal area, the motor trigeminal nucleus, the motor trigeminal nucleus, the facial, hypoglossal and parabrachial nuclei and specific parts of the caudal parvocellular reticular formation, including nucleus linearis and the dorsal reticular nucleus of the medulla. The raphe nuclei, nucleus of the solitary tract, inferior olive, dorsal principal sensory, spinal trigeminal nuclei and gigantocellular reticular nucleus and the ventral reticular nucleus of the medulla received moderate projections. In general, the projections from the rostral parvocellular reticular formation were bilateral with an ipsilateral dominance. The dorsal motor vagus and the ambiguus nuclei were not labeled. It is concluded that the rostral parvocellular reticular formation participates in regulation of orofacial motor control and in neural networks for limbic control of metabolic homeostasis.

SELECTIVE EXPRESSION OF NEUROTROPHIN-3 MESSENGER-RNA IN MUSCLE-SPINDLES OF THE RAT

The expression of neurotrophin-3 messenger RNA was studied by in situ hybridization in rat muscle spindles from the first embryonic stages of their formation until their mature appearance in adult animals. The first expression of neurotrophin-3 messenger RNA in developing muscles was observed at E19 in the firstly formed intrafusal fiber, the nuclear bag2 fiber. High levels of neurotrophin messenger RNA were found in the equatorial region of these intrafusal fibers in thin lines of cytoplasma around and between the packed-up nuclei. From E21 on, neurotrophin-3 messenger RNA was also present in the nuclear bag1 type intrafusal fiber. The expression of neurotrophin-3 messenger RNA in nuclear chain fibers, which were found in muscle spindles from day 6 after birth, was low and insignificant in comparison to the expression in the nuclear bag fibers. After completion of muscle spindle formation around the third week after birth, high levels of neurotrophin-3 messenger RNA remained present in the intrafusal fibers throughout life. During the entire period of muscle formation, examined from E15 on, as well as in mature muscles, no neurotrophin-3 messenger RNA could be detected in extrafusal fibers by in situ hybridization. The exclusive intramuscular expression of neurotrophin-3 messenger RNA in intrafusal fibers during development as well as in mature stages suggests the involvement of neurotrophin-3 in the formation and the maintenance of muscle spindles.

Neurotransmitters and neuropeptides within the mesencephalic trigeminal nucleus of the rat: An immunohistochemical analysis

In order to determine which neurotransmitters and neuropeptides are utilized by the neurons of the mesencephalic trigeminal nucleus and by the fibres making synaptic contact with these primary sensory cells, we have set up an immunohistochemical study using antibodies against 17 major neurotransmitters and neuropeptides in the rat. Apart from some intracellular immunostaining for glutamate, no immunoreactivity to any of the tested neurotransmitters and neuropeptides could be detected inside mesencephalic nucleus of the trigeminal nerve neurons. Our immunohistochemical observations indicate that mesencephalic nucleus of the trigeminal nerve neurons receive input from various nerve fibres that appear to utilize serotonin, GABA, dopamine, noradrenaline (and likely glutamate) as transmitters. The innervation appeared randomly distributed over all mesencephalic nucleus of the trigeminal nerve neurons. The presence of substance P, cholecystokinin, vasoactive intestinal polypeptide, bombesin/gastrin releasing peptide, [Leu]enkephalin and neuropeptide Y observed in some fibres that contact with mesencephalic nucleus of the trigeminal nerve neurons, presumably reflect the co-existence of these peptides with one of the neurotransmitters.

Ultrastructure of the Rat Mesencephalic Trigeminal Nucleus

The subcellular morphology of the mesencephalic trigeminal (Me5) nucleus in the rat was studied by transmission electron microscopy. Most neurons in the thin rostral as well as in the major caudal part of Me5 appeared as large (40-50 microns), round- to ovoid-shaped unipolar cells. A few neurons (estimated 5%) appeared to be multipolar, usually bipolar. The Me5 neurons had a large, round, centrally located nucleus, and their cytoplasm was characterized by a dense network of lamellar granular endoplasmic reticulum, an abundant Golgi apparatus, many mitochondria and neurofilaments suggesting very active cells with a high rate of synthesis and axoplasmatic transport. Numerous small spinous processes covered the surface of the Me5 neurons. Clustering of 2 or 3 cells was accomplished by maculae, i.e. zones of gap junctions and close cell appositions. Boutons contacting the soma of Me5 neurons and boutons contacting large and small dendrites were defined as axosomatic and axodendritic synapses, respectively. Four types of synaptic boutons were distinguished: (1) S boutons, with round vesicles and asymmetrical as well as symmetrical synapses, (2) F boutons, with pleomorphic admixture of flattened and spherical vesicles and asymmetrical synapses, (3) P boutons, which resembled the F-type boutons but contained predominantly spherical vesicles and symmetrical synapses, and (4) G boutons, characterized by a heterogeneous population of vesicles. This description of the Me5 nucleus is particularly useful for future studies that attempt to correlate the structure of a particular synapse with its function.

ORIGIN, DISTRIBUTION AND MORPHOLOGY OF SEROTONERGIC AFFERENTS TO THE MESENCEPHALIC TRIGEMINAL NUCLEUS OF THE RAT

We have studied the localization, the morphology and sources of serotonergic input on the primary afferent neurons in the mesencephalic trigeminal nucleus (Me5) of the rat with light and electronmicroscopy immunocytochemistry and with anterograde and retrograde neuroanatomical tract tracing methods. Me5 neurons were found to receive a serotonergic input that is part of a serotonergic fibre plexus extending over the neighbouring parabrachial nucleus and locus coeruleus. These serotonergic afferents originate predominantly from serotonergic cells in the dorsal raphe nucleus.

DOPAMINERGIC AFFERENTS TO THE MESENCEPHALIC TRIGEMINAL NUCLEUS OF THE RAT - A LIGHT AND ELECTRON-MICROSCOPE IMMUNOCYTOCHEMISTRY STUDY

The localization and sources of dopaminergic projections on the primary afferent neurons in the mesencephalic trigeminal nucleus (Me5) of the rat were studied using light and electron microscopic immunocytochemical staining techniques combined with anterograde and retrograde neuroanatomical tract tracing methods. Me5 neurons were found to receive a dopaminergic input that is part of a dopaminergic fibre plexus extending over the neighbouring nucleus parabrachialis and locus coeruleus. These dopaminergic afferents originate from the substantia nigra, the ventral tegmental area and the medial hypothalamus.

Distribution of Synaptic Boutons in the Mesencephalic Trigeminal Nucleus of the Rat – A Quantitative Electron-Microscopical Study

The distribution of synapses and synaptic bouton types in the mesencephalic trigeminal (Me5) nucleus was examined in a quantitative electron-microscopical study. Of 588 terminal boutons that were counted in the compact caudal part of the Me5 nucleus, less than 8% formed synapses on the somata of the predominantly unipolar Me5 neurons. About 79% formed synapses on fibres located between the Me5 somata, while about 13% of the vesicle-containing terminals had no clear synaptic specialization. All of these non-synaptic terminals were G type boutons, with pleomorphic and large characteristic dense-core vesicles. Approximately 60% of the axosomatic synapses were of the S type, containing spherical vesicles and an asymmetrical or symmetrical synaptic specialization. About 20, respectively 15% of the axosomatic synapses, were of the F, respectively P type; both are symmetrical synapse types containing either a majority of flat or pleomorphic vesicles. Less than 10% of the axosomatic synapses were of the G type. Although some proportional differences were noted, an almost similar bouton type distribution pattern was found for the axodendritic synapses suggesting that the axosomatic and axodendritic synapses in the Me5 nucleus are part of the same afferent fibre plexus covering the Me5 nucleus.

Ultrastructural localisation of intramuscular expression of BDNF mRNA by silver-gold intensified non-radioactive in situ hybridisation.

Abstract. A non-radioactive in situ hybridisation method is described for the detection of low intramuscular levels of brain-derived neurotrophic factor (BDNF) mRNA at the electron microscope level. Application of high-grade silver-gold intensification of the diaminobenzidine end product of in situ hybridisation revealed that in adult rat muscle the constitutive expression of muscular BDNF is confined to the myofibres; satellite cells, Schwann cells, endothelial cells, fibroblasts or axons do not appear to contribute to BDNF production in normal muscle. Although muscular BDNF is a neurotrophic factor for innervating motoneurons and supposedly released only at the motor endplates, the production of BDNF mRNA appears to occur along the entire length of the myofibres and is not confined to nuclei in the postsynaptic regions.

Neurotrophin-3 mRNA expression in rat intrafusal muscle fibres after denervation and reinnervation

We have studied the regulation of the expression of neurotrophin-3 (NT-3) mRNA in neonatal and adult rat muscle spindles after denervation and after denervation followed by reinnervation. Denervation of the intrafusal fibres did not result in an upregulation of the NT-3 mRNA expression but decreased this expression below the detection limit of the in situ hybridization method. Reinnervation of intrafusal fibres restored the NT-3 mRNA expression. The results suggest that the expression of NT-3 mRNA in postnatal muscle spindles is controlled by neuronal factors. The intrafusal fibre derived NT-3 may act as an instructive, feedback messenger for innervating neurons and may play a role in stabilizing and maintaining functional neuromuscular connections.

CONTRALATERAL PROJECTIONS OF CELLS IN THE MOTOR TRIGEMINAL NUCLEUS OF THE RAT

With horseradish peroxidase and Phaseolus vulgaris lectin as tracers, a direct connection between the jaw closing parts of the ipsi- and contralateral motor trigeminal (Mo5) nuclei of the rat is shown. The contralateral projecting cells in Mo5 were small (18 X 11 microns) ovoid and bipolar. It is speculated that these contralateral projecting cells in Mo5 are interneurons that are involved in the coordination of the bilateral activity of jaw closing motoneurons during orofacial motor behavior.