J. I. Lehtosalo

Distribution of adrenergic nerves in the lacrimal glands of guinea-pig and rat

The distribution of adrenergic nerves in the ex- and intraorbital lacrimal glands of guinea-pig and rat was studied using the sucrose-potassium phosphate-glyoxylic acid (SPG) technique. Blood vessels and secretory acini of guinea-pig lacrimal glands were demonstrated as having a rich adrenergic innervation. Adrenergic fibers in the rat were, however, much more sparse, and most of them were seen in association with glandular blood vessels, with only a few being found between secretory acini. Pretreatment of animal with a monoamine oxidase inhibitor and L-dopa did not change the morphological distribution of catecholamine fluorescent fibers, although the treatment improved the fluorescence, especially in the rat. Extirpation of the ipsilateral superior cervical ganglion eliminated all the fluorescent fibers in both normal and pretreated animals. The presence of adrenergic innervation of the lacrimal glands, especially in close connection with secretory acini, supports the theory that catecholamine-containing nerves play a role in the regulation of lacrimal secretion.

Biochemical and immunohistochemical determination of 5-hydroxytryptamine located in mast cells in the trigeminal ganglion of the rat and guinea pig

SummaryThe presence of 5-hydroxytryptamine (5-HT), as well as its precursor (5-HTP) and metabolite (5-HIAA), were biochemically determinated in the trigeminal ganglion of the guinea pig and rat. The distribution of 5-HT in the ganglion and in its posterior root was studied using both indirect immunofluorescence and the peroxidase-antiperoxidase method. In order to increase the possible 5-HT content of primary sensory neurons for subsequent immunohistochemical visualization, animals were first treated with nialamide, an inhibitor of monoamine oxidase, and then loaded with l-tryptophan. Another group of animals received colchicine to inhibit intra-axonal transport of transmitter substances. However, even combined use of loading and colchicine treatment did not reveal 5-HT immunoreactivity in ganglion cells.The only source of 5-HT immunoreactivity in the trigeminal ganglion and its posterior root was mast cells. These cells were located around the ganglion in adjacent leptomeningeal and connective tissues, as well as between the ganglion cells and nerve fibers. Only occasionally were mast cells found in the posterior root of the ganglion.

Substance P-like immunoreactive trigeminal ganglion cells supplying the cornea

SummaryTrigeminal ganglion cells supplying the cornea were traced with intra-axonally transported horseradish peroxidase and, subsequently studied for the presence of substance P-like immunoreactivity. Approximately 0%–30% of trigeminal ganglion cells contained immunoreactive substance P. These cells were of a small size (15–50 μm in diameter) and were distributed throughout the ganglion. The ganglion cells supplying the cornea were of a relatively small size as well but were confined to the anteromedial part of the ganglion. Some of these cells were found to contain immunoreactive substance P.

Is Substance P the Neurotransmitter in the Vestibular End Organs

The vestibular end organs of the rabbit were fixed intracardially with phosphate buffered 4% formaldehyde, sectioned for 15 micron cryostat sections and incubated with monoclonal substance P (SP) antibody. Specific SP-like immunoreactivity (SPLI) was observed within and below the sensory epithelia of both utricular and saccular maculae and ampullary cristae. Particularly strong SPLI was seen in the maculae which showed many SPLI-positive axons in the subepithelial space and a network of SPLI-positive structures in the basal zones of the sensory epithelium. The calyceal terminals of type I hair cells showed regularly SPLI. The possibility of a transmitter or modulator role for SP in the vestibular function is suggested.

Immunohistochemical localization of unique enkephalin sequences contained in preproenkephalin A in the guinea pig cochlea.

The guinea pig cochlea was studied for the presence of immunoreactivities to the four unique enkephalin sequences contained in the preproenkephalin A. The antisera to Met-enkephalin-Arg6-Phe7 and Met-enkephalin-Arg6-Gly7-Leu8 were used as highly specific markers for the preproenkephalin A. Contrary to Met-enkephalin and Leu-enkephalin, also included in the present study, these two peptide sequences are not contained in pro-opiomelanocortin or preproenkephalin B (prodynorphin). All the four different antisera showed identical localization for the four peptide sequences, suggesting their coexistence in the same nervous pathway. Specific immunofluorescence was found in intraganglionic spiral bundle, inner spiral bundle, tunnel spiral bundle and in association with inner hair cells. The nerve fibers were thin and varicose, suggesting that most, if not all, of them were unmyelinated. Their localization indicates that the unique enkephalin sequences contained in preproenkephalin A are present in the cochlear efferent pathway to the inner hair cell region.

Nerve fibers showing immunoreactivities for proenkephalin A-derived peptides in the lacrimal glands of the guinea pig

The extra- and intraorbital lacrimal glands of guinea pigs were studied for the presence and distribution of enkephalin-like immunoractive nerve fibers. Four specific antisera against the four different enkephalin sequences contained in pre-proenkephalin A (Met-Enk, Met-Enk-Arg-Phe, Met-Enk-Arg-Gly-Leu, and Leu-Enk) were used. All of these immunoreactivities were identically distributed in varicose nerve fibers in both the extra- and intraorbital lacrimal glands. These fibers densely surrounded the glandular acini and also innervated the secretory ducts. Sympathetic denervation had no effect on these nerve fibers. The results suggest that enkephalins derived from proenkephalin A may play a role in the nervous control of the lacrimal secretion.

Substance P-like immunoreactivity in the trigeminal ganglion

SummaryThe trigeminal ganglion of rat and guinea pig was studied for the presence of immunoreactive substance-P using fluorescence, light and electronmicroscopy. In untreated animals substance P containing cells, with a diameter of 15 to 50 μm, were distributed throughout the ganglion and comprised 10–30% of all ganglion cells. Colchicine, injected intraventricularily to inhibit intra-axonal transport, had no effect on the number of substance P cells; but when the drug was injected directly into the posterior root of the ganglion, the proprotion of these cells increased to as much as 50%. In the electron microscope, immunoreactive substance-P was confined to ganglion cells classified as B type according to the arrangement of subcellular organelles, and to unmyelinated nerve fibers. Subcellularily the immunoreactivity appeared in cytoplasmic vesicles, as well as dispersed in the nerve fibers and the perikarya of neurons. The great number of substance P immunoreactive ganglion cells suggests that they do not comprise a well defined subpopulation of the B-cells.However, the immunoreactivity was restricted to a distinct ultrastructural type of neurons with unmyelinated nerve fibers, suggesting that they also may share some distinct functions.

Sensory supply of the anterior uvea: A light and electron microscope study

SummaryWe have studied the ultrastructural features of sensory nerve fibers in the ciliary body and the iris and their parent trigeminal ganglion cells, using intra-axonally transported horseradish peroxidase as a tracer. Unmyelinated nerve fibers of ipsilateral trigeminal origin were found in stroma of both the ciliary body and the iris. Most appeared in bundles of nerve fibers; but some were found singly. Two distinct types of varicosities were found, one containing both a few irregularly shaped vesicles and mitochondria, the other containing mitochondria only. The thin profiles of the axons between these varicosities or endings contained neurotubules and filaments. The trigeminal ganglion cells supplying the anterior eye were of a relatively small size (15–50 μm in diameter) and confined to the anteromedial part of the ipsilateral ganglion. These cells could not be classified according to the arrangement of their subcellular organelles. The morphological characteristics of the sensory nerve fibers and ganglion cells supplying the anterior uvea are consistent with the view that pain may be the predominant sensation mediated by these neurons.

Vasoactive intestinal polypeptide (VIP)-immunoreactive nerve fibers in the anterior uvea of the guinea pig.

The presence of vasoactive intestinal polypeptide (VIP)-like immunoreactivity, especially in the anterior part of the guinea pig uvea, was studied using an indirect immunofluorescence technique. In the ciliary body, thin varicose VIP fibers displaying VIP immunoreactivity were observed in the ciliary muscle and in the ciliary processes. The iris was virtually free of VIP fibers. In the choroid, VIP-immunoreactive fibers were seen mainly in close association with the choroidal blood vessels. The histochemical localization of VIP-immunoreactive fibers in the anterior uvea suggests that the physiological role of neuronal VIP in the uvea is not only associated with the regulation of the blood flow of the choroid but also with the functions of ciliary muscle and ciliary epithelium.

Recovery of the blood-aqueous barrier after topical chemical irritation in the rabbit eye.

An acute irritative response in the rabbit eye, taking the form of a rise in the intraocular pressure (IOP), miosis, and breakdown of the blood-aqueous barrier, was elicited by topical application of 1% neutral formaldehyde. The peak rise in IOP was 22.1 ± 2.5 mmHg and occurred within 14.2 ± 1.9 min, after which IOP returned to normal values in 45.4 ± 5.2 min. An increased amount of i.v.-injected Evans Blue was found in the aqueous humour when injected 15 min after the irritation (119.0 ± 21.2 μg/ml, compared with 2.1 ± 1.4 μg/ml in intact eyes;P < 0.001), and the protein concentration in the aqueous humour was also increased, to 11.3 ±1.4 mg/ml (P < 0.001). When Evans Blue was injected 60 min after the irritation, no statistically significant difference was found from normal in the amounts (13.7 ± 7.8 μg/ml) in the aqueous humour, although the protein concentration in the aqueous humour was again found to be elevated (7.9 ± 1.6 mg/ml,P < 0.01). Histologically, the Evans Blue was shown to leak through both the ciliary and the iris barriers at 15 min. At 60 min only occasional Evans Blue leakage was demonstrated between the nonpigmented epithelial cells and only minor fluorescence was seen in iris stroma. The histological findings were in good agreement with Evans Blue analyses in the aqueous humour. The miosis lasted over 60 min in the formaldehyde-treated eyes. After sympathectomy the rise in IOP was more rapid at the beginning of the response, suggesting greater sensitivity of the sympathectomized eyes to the irritative stimuli. However, the miosis had recovered by 60 min after irritation in the sympathectomized eyes. In the contralateral eyes changes in IOP and in the integrity of the blood-aqueous barrier were observed. The present results suggest that the breakdown of the blood-aqueous barrier after sensory nerve stimulation is transient and recovery rapid.