Presence of neuronal nitric oxide synthase in autonomic and sensory ganglion neurons innervating the lacrimal glands of the cat: an immunofluorescent and retrograde tracer double-labeling study.

It is generally considered that parasympathetic postganglionic nerve fibers innervating the lacrimal gland (LG) arise from the pterygopalatine ganglion (PPG), while sympathetic and sensory innervations arise from the superior cervical ganglion (SCG) and trigeminal ganglion (TG), respectively. Recently, we reported for the first time that the parasympathetic innervation of the cat LG was also provided by the otic ganglion (OG) and ciliary ganglion (CG), and that the sensory innervation was also provided by the superior vagal ganglion (SVG) and superior glossopharyngeal ganglion (SGG). To determine if nitric oxide (NO) is a neurotransmitter of the autonomic and sensory neurons innervating the LG, we injected the cholera toxin B subunit (CTB) as a retrograde tracer into the cat LG, and used double-labeling fluorescent immunohistochemistry for CTB and nitric oxide synthase (NOS). We found that NOS-/CTB-immunofluorescent double-labeled perikarya were localized in the PPG, OG, TG, SVG and SGG, but not in the CG and SCG. The highest numbers of NOS-/CTB-immunofluorescent double-labeled neurons were found in the PPG and TG. In addition, we examined the presence of nitrergic nerve fibers in the LG using NADPH-d histochemistry and found that a large amount of NADPH-d-stained nerve fibers were distributed around the glandular acini and in the walls of glandular ducts and blood vessels. This study provides the first direct evidence showing that NO may act as a neurotransmitter or modulator involved in the parasympathetic and sensory regulation of lacrimal secretion and blood circulation, but may not be implicated in the sympathetic control of LG activities, and that nitrergic nerve fibers in the LG arise mainly from parasympathetic postganglionic neurons in the PPG and sensory neurons in the TG. The present results suggest that NO plays an important role in the regulation of LG activities.

Three novel neural pathways to the lacrimal glands of the cat: an investigation with cholera toxin B subunit as a retrograde tracer.

The distribution of ganglion neurons innervating the lacrimal gland (LG) was investigated following injection of cholera toxin B subunit into the LG of the cat. We report the first evidence that the otic ganglion (OG), and superior vagal and glossopharyngeal ganglia are also the sources of innervation of the LG. LG-innervating neurons in the pterygopalatine ganglion and the OG could be divided into two subpopulations: small and large neurons. They may mediate the vasodilatation and secretion, respectively.

Morphological distinction between vasodilator and secretomotor neurons in the pterygopalatine ganglion of the cat.

We investigated whether vasodilator and secretomotor ganglion neurons are morphologically distinguishable from each other in the parasympathetic ganglion of the cat. When Cholera toxin B subunit, a retrograde tracer, was injected into the palatine gland, both large and small ganglion neurons were retrogradely labeled in the pterygopalatine ganglion. On the other hand, when the tracer was injected into gland-free areas (the upper gingiva or epidural space), all neurons labeled in the ganglion were small in size. Thus, it was assumed that small and large neurons labeled in the ganglion represented, respectively, vasomotor and secretomotor and neurons [corrected].

Facial nerve parasympathetic preganglionic afferents to the accessory otic ganglia by way of the chorda tympani nerve in the cat.

The distribution of accessory otic ganglia and connections between the ganglia and the chorda tympani nerve were investigated in the cat in order to determine the parasympathetic preganglionic facial nerve afferents to the otic ganglia using whole mount acetylthiocholinesterase (WATChE) histochemistry. The otic ganglia consist of a single main prominent ganglion and many small accessory ganglia lying on a plexus around the origins of the branches of the mandibular nerve and near the junction of the chorda tympani nerve and lingual nerve. In cell analysis of Nissl-stained preparations, the neurons composing the accessory otic ganglia were morphologically similar to the main otic ganglion neurons. Connecting branches from the chorda tympani nerve to the peripherally located accessory otic ganglia were found and they were not stained by WATChE histochemistry. WATChE-positive connecting branches from the ganglia to the inferior alveolar, lingual, and mylohyoid nerves were also found in the same preparations. The WATChE histochemistry on various autonomic nervous tissues revealed that autonomic postganglionic nerve fibers are selectively stained darkly and that preganglionic fibers remain unstained. Therefore, it is considered that the WATChE-negative connections from the chorda tympani nerve consist chiefly of autonomic preganglionic fibers, whereas the WATChE-positive connections to the branches of the mandibular nerve are mainly postganglionic fibers. This suggests that some of the facial nerve parasympathetic preganglionic fibers in the chorda tympani nerve are mediated in the accessory otic ganglia and then join the branches of the mandibular nerve to supply the target mandibular tissues.

Autonomic and sensory innervation of cat molar gland and blood vessels in the lower lip, gingiva and cheek.

Innervation of the molar gland and blood vessels in the lower lip, gingiva and cheek mucous membrane was investigated in the cat with the aid of whole mount acetylthiocholinesterase (WATChE) histochemistry and retrograde neuronal tracing methods with horseradish peroxidase (HRP) and HRP-conjugated wheat germ agglutinin (WGA-HRP). The molar gland was found to be supplied from the buccal nerve and branches of the mylohyoid nerve on the basis of microdissection of WATChE-stained mandibular preparations under a dissecting microscope. The rostral half of the lower lip-gingiva was innervated by mental branches from the inferior alveolar nerve. The caudal half of the lower lip-gingiva and cheek mucous membrane were observed to be supplied from the buccal nerve. Following injections of HRP/WGA-HRP into the molar gland, lower lip-gingiva and cheek, many retrogradely labeled ganglion neurons were observed in the ipsilateral main and accessory otic ganglia, superior cervical ganglion and mandibular division of the trigeminal ganglion. In the pterygopalatine ganglion, a small number of positive neurons were found, but in a few cases in which the injected tracer was restricted to the lower lip-gingiva and anterior half of the molar gland, labeled neurons were not detected in the main ganglion nor in its accessory microganglia. These findings indicate that the cat molar gland receives a postganglionic parasympathetic supply from the otic ganglia, postganglionic sympathetic input from the superior cervical ganglion and sensory innervation from the trigeminal ganglion by way of the buccal nerve and mylohyoid nerve. Vessels in the rostral half of the lower lip-gingiva receive the same inputs from the inferior alveolar nerve, and vessels in the caudal half receive inputs from the buccal nerve. The vessels in the cheek mucous membrane receive dual parasympathetic supplies from the otic ganglia and the pterygopalatine ganglion by way of the buccal nerve.

NADPH diaphorase neurones are evenly distributed throughout cat neocortex irrespective of functional specialization of each region.

Using NADPH diaphorase histochemistry as a marker for nitric oxide synthase we investigated the possible sites of nitric oxide synthesis in cat cerebral neocortex. Intensely stained neurones were found mainly in the deep layers of the neocortex and underlying medulla. Virtually all neurones in the cerebral medulla were NADPH diaphorase positive. The density of diaphorase neurones was estimated in the cortex/medulla border zones of each neocortical gyrus. Diaphorase neurones were evenly distributed throughout the neocortex and no significant statistical difference between gyri was observed. These findings indicate that the density of diaphorase neurones is irrespective of functional specialization of each region and are more in line with the hypothesis that NADPH diaphorase neurones are involved in the control of local cortical blood flow.

Localization of preganglionic neurons of the accessory ciliary ganglion in the midbrain: HRP and WGA-HRP studies in the cat.

Localization of preganglionic neurons of the accessory ciliary ganglion (ACG), including ectopic intraocular ganglion cells, was investigated in the cat with the aid of horseradish peroxidase (HRP) and HRP-conjugated wheat germ agglutinin (WGA-HRP) methods. When HRP or WGA-HRP was injected into the anterior and posterior chambers of the eye, no retrogradely labeled cells were found in the visceral oculomotor nuclei, although most neurons of the ACG and the main ciliary ganglion (CG) were intensely labeled. When a microsyringe needle was inserted into the ciliary body, the tracer diffused into the suprachoroid lamina and the intraocular ganglion cells, and a small number of labeled neurons appeared in the midplane between each side of the somatic oculomotor nuclei. After injection into the ACG, many labeled neurons were observed in the anteromedian nucleus, Edinger-Westphal nucleus, and midplane between the somatic oculomotor nuclei, their ventral continuations of the ventral tegmental area, and the periaqueductal gray. HRP/WGA-HRP injection into the CG labeled cells in all these areas and in the lateral border zones of the anteromedian, Edinger-Westphal and somatic oculomotor nuclei, and their ventral continuations of the ventral tegmental area. These findings indicate that the visceral oculomotor neurons which project to the ACG tend to be located more medially than those to the CG.

Oculomotor parasympathetic pathway to the accessory ciliary ganglion bypassing the main ciliary ganglion by way of the trigeminal nerve.

When an HRP or WGA-HRP solution was injected into the rostral midbrain including the oculomotor visceral nuclei, densely distributed HRP/WGA-HRP-positive granules were observed around the ganglion neurons in the accessory ciliary ganglion (ACG) and ectopic neurons in the communicating branch from the long ciliary nerve to the ACG. The same injections labeled fibers within the communicating branch as well as the fibers between the ACG and the main ciliary ganglion (CG). These findings indicate that some oculomotor parasympathetic preganglionic fibers reach the ACG bypassing the CG by way of the trigeminal nerve.

Comparative anatomy of the accessory ciliary ganglion in mammals.

The orbits of 13 mammalian species (pig, sika deer, domestic sheep, horse, cat, fox, racoon dog, marten, rat, rabbit, crab-eating macaque, japanese macaque and man) were stained with silver nitrate and dissected under a dissecting microscope with special attention to the presence and location of the accessory ciliary ganglion. Some preparations were stained with thionin and examined as whole-mounts in a transmission microscope. The accessory ciliary ganglion was present in all 13 species, although the number and degree of development varied greatly from species to species. The accessory ciliary ganglion could be readily differentiated from the main ciliary ganglion in the following respects: it was located on the short ciliary nerve, and it had no root derived directly from the inferior trunk of the oculomotor nerve and it never attaches to this nerve. In many species, ganglion cells were also scattered in the short ciliary nerves in the stained whole preparations. In a few species, there were one or more small ganglia on the nerve to the inferior oblique muscle.

Efferent connections of area 20 in the cat: HRP-WGA and autoradiographic studies.

Following injections of horseradish peroxidase-wheat germ agglutinin conjugate (HRP-WGA) and tritiated leucine into area 20 of the cat, terminal labeling was observed in visual areas 19, 21, the splenial visual area, the lateral suprasylvian area as well as in premotor, association and limbic related cerebral cortical regions. Labeled terminals in the subcortex were distributed in the caudate nucleus, the claustrum, the putamen, the anterior ventral nucleus, the intralaminar nuclei, the caudal division of the intermediate lateral nucleus, the lateralis posterior-pulvinar complex, the parvocellular C laminae of the dorsal lateral geniculate nucleus and the ventral lateral geniculate nucleus. In HRP-WGA preparations, retrogradely labeled somata were observed in these regions with the exception of certain subcortical structures. The projections are discussed with respect to the possible role area 20 plays in the cortical control of pupillary constriction.

Pupillo-motor areas in the rabbit visual cortex.

Pupillo-motor areas in the visual cortex of the rabbit were studied. Electrical stimulation of the rabbit visual cortex induced pupillary constriction or dilation. The low threshold area (0.5-1.0 mA) for pupillary dilation was in the form of a narrow band extending rostro-caudally along the boundary between the striate cortex (VI) and the occipital cortex (VII), where the binocular area of the visual field is represented. The pupillo-constrictory cortical points were also identified in the caudal part of the splenial gyrus (lateral part) and the pupillo-dilator points in the rostral part of the gyrus (lateral part).

Afferents to the cortical pupillo-constrictor areas of the cat, traced with HRP.

Afferents of the cortical pupillo-constrictor areas (PCAs) of the cat were studied using the horseradish peroxidase method. PCAs receive heavy cortical inputs from areas 7, 19, 21, the lateral suprasylvian area, the splenial visual area, and subcortically from the claustrum, the intralaminar nuclei, the pulvinar-lateral posterior nuclear complex.