Involvement of the pelvic plexus and the suprarenal ganglia in the neuropeptide Y (NPY) innervation of the cervix and the uterus of the rat

The involvement of the pelvic plexus and suprarenal ganglia in the neuropeptide Y (NPY) innervation of the genital tract was studied in the female rat by means of denervation experiments and retrograde tracing studies. Removal of the paracervical ganglia caused a significant decrease of the NPY-immunoreactive nerve density and NPY concentration in the lower part of the genital tract: cervix, uterine body and lower part of the uterine horn. The decrease in NPY concentration in these three regions was more pronounced after lesion of the pelvic plexus. Lesion of the ovarian nerve plexus caused a depletion in the NPY-immunoreactive nerve fibres and a decrease in NPY concentration in the upper part of the uterine horn. Pelvic nerve section, inferior mesenteric ganglia excision and superior ovarian nerve section had no effect on the NPY innervation in the genital tract. Injection of fluorogold into the cervix and lower part of the uterus combined with immunohistochemistry revealed that 87.5% of labelled neurons in the pelvic plexus were NPY-immunoreactive. Following injection of fluorogold into the upper part of the uterus, 92% of labelled neurons in the suprarenal ganglia were NPY-immunoreactive. Treatment with 6-hydroxydopamine revealed that the NPY-immunoreactive nerve fibres were non-noradrenergic in the cervix, but were noradrenergic in the upper part of the uterus. In the uterine body and lower part of the uterine horn, both noradrenergic and non-noradrenergic NPY-immunoreactive nerve fibres were observed. These data demonstrate the major contribution of pelvic plexus neurons in the non-noradrenergic NPY innervation of the lower part of the genital tract, and the involvement of the suprarenal ganglia in the noradrenergic NPY innervation of the upper part of the uterus via the ovarian nerve plexus.     

A quantitative assessment of innervation in the conduction system of the calf heart

BACKGROUND: The aim of the present investigation was to determine the relative distribution of autonomic and sensory nerves in the cardiac conduction tissues of calves. METHODS: A quantitative immunohistochemical and histochemical technique was adopted. RESULTS: Immunoreactivity to the general neuronal marker protein gene product 9.5 (PGP 9.5) demonstrated that all regions of the conduction system possessed a higher relative density of total nerves when compared with the surrounding myocardial tissues. Unlike myocardial innervation, the conduction system did not display an atrial-to-ventricular gradient in nerve density. PGP 9.5-immunoreactive nerve trunks and varicose nerve fibres were more numerous in the transitional atrioventricular node and the penetrating atrioventricular bundle than in either the sinus node, compact atrioventricular node, or bundle branches. The Purkinje network of the ventricular conduction tissues possessed a rich supply of PGP 9.5-immunoreactive nerve trunks and varicose nerve fibres. Acetylcholinesterase (AChE)-positive nerves were the main subtype identified in the sinus and atrioventricular nodes and in the ventricular conduction tissues, representing 50-80% of the area occupied by PGP 9.5-immunoreactive nerves. The compact atrioventricular node possessed AChE-positive and tyrosine hydroxylase (TH)-immunoreactive nerves in similar proportions (45%), although, in general, TH-immunoreactive nerves had a lower relative nerve density than AChE-positive nerves. Neuropeptide Y (NPY)-immunoreactive nerves represented the main peptide-containing subpopulation and occurred throughout the conduction system, displaying a similar pattern of distribution and relative density to those demonstrating TH immunoreactivity. Nerve fibres immunoreactive for somatostatin, vasoactive intestinal polypeptide, substance P, and calcitonin gene-related peptide formed relatively minor subpopulations. CONCLUSIONS: The general innervation of the bovine conduction tissues exhibits significant regional variation. Throughout all regions of the conduction system, AChE-positive nerve represented the dominant subtype when compared with TH- and NPY-immunoreactive nerves. The distribution and relative density of nerve subtypes in the tissues of the bovine conduction system are similar to those observed in man, whereas differences were observed in other regions, such as the atrioventricular bundle and bundle branches. This finding must be considered by those making interspecies comparisons.     

Peripheral sympathetic innervation of the deep pineal gland of the golden hamster

Both the superficial and deep pineal components of the intact hamster contain a rich network of green to yellow-green fluorescent nerve fibres. After either superior cervical ganglionectomy or after transection of the nervi conarii the majority of the fluorescing fibres disappeared from both the superficial and deep pineal masses. Although the deep pineal remained intact after surgical removal of the superficial pineal, it was devoid of any green or yellow-green fluorescent fibres.     

Neuronal projections from the mesencephalic raphe nuclear complex to the suprachiasmatic nucleus and the deep pineal gland of the golden hamster (Mesocricetus auratus)

Neuronal projections from the mesencephalic raphe system to the suprachiasmatic nucleus and the pineal complex were mapped in this study of the golden hamster, by use of the anterograde tracer Phaseolus vulgaris-leucoagglutinin and the retrograde tracer cholera toxin subunit B. From the median raphe nucleus, a rostral projection ascended in the ventral part of the mesencephalon to continue in the medial forebrain bundle of the forebrain. Nerve fibres from this bundle innervated the ventral and medial parts of the suprachiasmatic nucleus. At the level of the interpeduncular nucleus of the mesencephalon, fibres of the ventral bundle bent dorsally to reach the epithalamic area and to continue in the forebrain in a periventricular position. Some of these fibres innervated the dorsal tip of the suprachiasmatic nucleus. The dorsal raphe nucleus was the origin of a nerve fibre bundle, located in the periaqueductal gray of the mesencephalon, innervating the deep pineal gland and pineal stalk. Injection of cholera toxin B into the suprachiasmatic nucleus labelled cells in the median raphe. Combination of the retrograde tracing from the suprachiasmatic nucleus and serotonin transmitter immunohistochemistry showed that some of the cholera toxin B-immunoreactive nerve cells also contained serotonin. Thus, this study of the golden hamster shows a serotonergic projection from the median raphe nucleus to the suprachiasmatic nucleus and a projection from the dorsal raphe nucleus to the deep pineal gland supporting physiological indications of an influence of serotonin on the photoreceptive circadian system of the brain.     

Innervation of the pancreas of the one-humped camel (Camelus dromedarius) by neuropeptide-Y, galanin, calcitonin-gene-related-peptide, atrial natriuretic peptide and cholecystokinin

The distribution of neuropeptide-Y (NPY), galanin (GAL), cholecystokinin-8 (CCK-8), atrial natriuretic peptide (ANP) and calcitonin gene-related peptide (CGRP) in the pancreas of the camel was investigated using immunohistochemical techniques. NPY-immunoreactive neurons were observed in the pancreatic ganglia and also in the interacinar regions of the exocrine pancreas. NPY was discernible in fine varicose nerve fibres ending on NPY-negative cells and in the walls of blood vessels. ANP immunoreactivity was observed in nerve fibres situated on the basolateral surfaces of the acinar cells. CCK-8, GAL and CGRP immunoreactivity were observed in neurons and varicose nerve fibres in the walls of blood vessels. Of all the neuropeptides investigated, only NPY appeared to be densely distributed in the pancreas of the camel. It is concluded that the pattern of distribution of these neuropeptides in the camel pancreas is similar to those observed in the pancreata of other mammals.     

Vasoactive intestinal polypeptide (VIP)--immunoreactive nerve fibres in the mammary gland of the pig

Immunohistochemical studies have been performed to investigate the coexistence of VIP with dopamine-beta-hydroxylase (D(beta)H), vesicular acetylcholine transporter (VAChT), somatostatin (SOM) or neuropeptyd Y (NPY) within nerve fibres supplying the immature mammary gland in the pig. Generally, a moderate number of the VIP-immunoreactive (VIP-IR) nerve fibres were located in the nipple and parenchyma of the gland. VIP-IR fibres surrounded smooth muscle cells (SMC), blood vessels (BV) and lactiferous ducts (LD). Double-labelling immunohistochemistry revealed that some of VIP-IR nerve fibres also contained immunoreactivity to D(beta)H. VIP/D(beta)H-IR nerves were associated with BV and SMC and single fibres were observed around the LD in both nipple and parenchyma of the gland. VIP/VAChT-IR nerve fibres were not observed. The majority of VIP-IR fibres associated with SMC were also SOM-IR. Less numerous VIP/SOM-IR fibres supplied the BV and were located around the LD of the gland. A small number of VIP-IR nerves also displayed immunoreactivity to NPY. VIP/NPY-IR nerve fibres supplied the BV of the gland.     

Pressure-independent inhibition of sympathetic activity by noradrenaline: role of baroreceptor C fibres

The effect of i.v. infusion of noradrenaline on activity in the renal sympathetic nerve was studied in rabbits anesthetized with chloralose and urethane. Noradrenaline (3--8 microgram/kg-min) initially increased mean arterial pressure 20--40 mmHg and consequently reduced renal nerve activity. However, studies over a wide range of pressures--obtained by changing the blood volume, revealed that noradrenaline after a few minutes had induced a pressure-independent reduction of sympathetic discharge. The effect disappeared with baroreceptor denervation. An unchanged relationship between arterial pressure and integrated activity in the whole left aortic nerve (which is largely a measure of activity in A fibres) suggested that the sympathetic depression was due to excitation of aortic nerve C fibres. This conclusion was supported by studies of sympathetic responses to selective stimulation of aortic nerve A and C fibres at equal pressures before and during infusion of noradrenaline. Compared to the reflex activity from A fibres, C fibre stimulation was invariably less effective in suppressing renal nerve activity during the infusion. Our studies indicate that noradrenaline may effect a negative feedback control of sympathetic discharge through activation of baroreceptor C fibres.     

Striatal neuropeptide Y neurones are not a target for thalamic afferent fibres

This study examined at the ultrastructural level the putative relationships between afferent fibres coming from the parafascicular nucleus of the thalamus and neuropeptide Y (NPY)-containing neurones in the rat striatum. Experiments used a combination of anterograde transport of the biotin dextran amine to label the thalamo-striatal pathway and immunogold labelling to reveal the NPY-containing neurones at the electron microscopic level. Examination of sections from three animals failed to demonstrate thalamic terminals in synaptic contact with NPY-immunoreactive dendrites or cell bodies, although both types of labelled elements were frequently involved in synaptic complex with unlabelled profiles. These results strongly suggest that striatal NPY interneurones are not under the direct influence of the main component of the thalamo-striatal system.     

Permeability of the pineal organ of the golden hamster (Mesocricetus auratus) to HRP with special reference to different types of blood capillaries

The pineal organ of the golden hamster consists of deep and superficial portions which are connected to each other by a stalk. The permeability of capillaries for horseradish peroxidase (HRP) injected intravenously was examined in sections of entire portions of the gland that cut either along coronal or sagittal planes. Two distinct portions of the parenchyma, i.e., dorsal major and ventral minor ones, were found in the superficial gland. Most of the capillaries in the dorsal portion were of the continuous type of endothelium, whereas those in the ventral portion were fenestrated. In the dorsal portion, HRP readily crossed the endothelium, permeated the basal lamina, flowed into the perivascular connective tissue space and intruded into the intercellular clefts of the parenchyma. In contrast, HRP was not found to penetrate through the endothelium of the capillaries in the ventral portion to reach the perivascular area, thereby leaving the intercellular clefts of the parenchyma free of HRP. In the deep gland the capillaries were exclusively of the nonfenestrated type. Intravenously injected HRP was prevented from crossing the endothelium by the tight junction. In some areas, HRP penetrated through the capillaries in the pia mater, and crossed the outer limiting membrane to reach the intercellular clefts of the parenchyma and the basal lamina of the capillaries in the peripheral region of the deep pineal gland. The junctions between endothelial cells were not penetrated by HRP. The observations indicate that the type of capillary, absence of perivascular spaces, and permeability in the deep pineal are all similar to these factors in the general brain tissue; they differ from these in the superficial pineal gland, in which the dorsal portion shows characteristics found in other endocrine glands, but the ventral zone exhibits a unique situations: the presence of a blood-pineal barrier with a pericapillary connective tissue area.     

Distribution of peptide-containing neurons in the developing rat right atrium, studied using immunofluorescence and confocal laser scanning

The developmental pattern and distribution of peptide-containing neurons in the rat heart right atrium has been studied by indirect immunofluorescence. Antibodies against neuropeptide Y (NPY), substance P (SP), and vasoactive intestinal polypeptide (VIP) were applied to whole-mount stretch preparations of the right atria from hearts of newborn to 40 day-old animals. NPY-like immunoreactivity (L1) was compared with the synaptic vesicle marker SV2 in double immunoincubation studies. The distribution of immunofluorescence was studied by confocal laser scanning microscopy. NPY-L1 and SP-L1 were present throughout the atria already at birth, in contrast to VIP-L1 that was observed at day 10. The postnatal changes of innervation were basically quantitative, with an increase in density of nerve fibres and number of varicosities, while the basic pattern of innervation was essentially established during the first 1-10 days. NPY- and SP-positive bundles of fibres appeared to enter the right atrium along the superior caval vein, having extrinsic origins. Nerve fibres with NPY-L1 colocalized in most nerve terminals with SV2-L1, and showed a developmental pattern similar to that observed for adrenergic neurons earlier. These NPY/SV2 positive fibres probably represent the extrinsic NPY innervation. In addition, NPY-L1 was identified in large intrinsic nerve cells bodies located near the atrioventricular (AV) region. Most of the VIP-L1 was observed in short nerve fibres originating in intrinsic VIP-positive cell bodies, but a few apparently extrinsic VIP-positive fibres were found, probably representing preganglionic parasympathetic neurons. SP in the atria was probably of extrinsic (sensory) origin and no nerve cell bodies with SP-L1 were detected. The results show that the peptidergic innervation in the developing rat right atrium involves both extrinsic and intrinsic peptidergic neurons which may participate in the regulation of neurotransmission in local neuronal circuits.     

Distribution and origin of peptide-containing nerve fibres in the rat and human mammary gland

The structures in the mammary gland involved in milk ejection have been investigated with regard to their relation to different types of peptidergic nerve fibres and their origin. Lactating rats were studied with immunohistochemistry focusing on the nipple, the parenchyma, the mammary blood vessels and the mammary nerve. The human mammary gland was also analysed. In the mammary gland from rat and human, nerve endings in the subepidermis, around smooth muscle cells in the nipple, in the connective tissue surrounding lactiferous ducts and alveoli in the nipple and in the parenchyma of the mammary gland showed immunoreactivity for calcitonin gene-related peptide, substance P, vasoactive intestinal polypeptide, peptide histidine isoleucine, neuropeptide Y, galanin and tyrosine hydroxylase, whereas dynorphin-positive nerve fibres could not be detected. The mammary nerve contained calcitonin gene-related peptide, vasoactive intestinal polypeptide, neuropeptide Y and tyrosine hydroxylase immunoreactivities; the adventitia of the mammary artery contained nerve fibres immunoreactive for neuropeptide Y and tyrosine hydroxylase, while vasoactive intestinal polypeptide-, peptide histidine isoleucine-, calcitonin gene-related peptide- and substance P-positive fibres were found in the tissue surrounding the artery. The wall of the mammary vein had nerve terminals immunoreactive for neuropeptide Y, tyrosine hydroxylase, calcitonin gene-related peptide and substance P. With the help of retrograde tracing using wheat germ agglutinin in combination with immunohistochemistry, projections of calcitonin gene-related peptide-immunoreactive cells in the dorsal root ganglia to the nipple were established. Neurons in the sympathetic stellate ganglion containing neuropeptide Y and tyrosine hydroxylase also projected to the mammary gland. Moreover retrogradely-labelled cells were found in the nodose ganglion, and they were vasoactive intestinal polypeptide-immunoreactive. These results demonstrate a rich distribution of different types of nerve fibres in structures of the mammary gland related to milk ejection. These nerve fibres and their peptides may be involved in the local control of milk ejection.     

Indications for a central innervation of the bovine pineal gland with substance P-immunoreactive nerve fibers

An antiserum, raised in rabbits, against substance P was used in an immunohistochemical investigation of the bovine pineal gland. A moderate innervation of all parts of the bovine pineal gland with substance P-immunoreactive nerve fibers was demonstrated. The immunoreactive nerve fibers were located throughout the pineal gland, both perivascularly, intraparenchymally, and with few fibers in the pineal capsule. Within the habenular nucleus, a large number of substance substance P-immunoreactive perikarya were present. From these perikarya processes extended towards the pineal stalk and gland. Some substance P-immunoreactive nerve fibers were located in the stria medullaris and in the posterior commissure. The anatomical location of the substance P-immunoreactive nerve fibers in the pineal gland and stalk strongly indicates that, in this species, substance P-immunoreactive pinealopetal nerve fibers originate from perikarya in the brain, probably from the medial habenular nucleus.     

Domestic violence

Phentolamine and propranolol (2 mg/kg i.p. of each) were administered to rats to be fed hard chow over a period of 80 min and to their fasting controls. The pharmacological elimination of the adrenoceptor mediated secretory drive on the parotid gland cells during feeding did not affect the gland content of the peptides substance P and vasoactive intestinal peptide localized to parasympathetic nerve fibres of the gland and thought to transmit secretory impulses. This is in contrast to the previously demonstrated effect of elimination of the parasympathetic acetylcholine-evoked secretion following administration of atropine, which resulted in reduced gland contents of these peptides.     

Region- and age-dependent variations of muscle fibre properties

The cytophotometric-morphometrical analysis of extensor digitorum longus and soleus muscles of 2.5 and 18 months old rats revealed regional and age-dependent differences in fibre type distribution, fibre area and fibre type related-enzyme activities which characterize contractility and metabolic profile. Variations along the longitudinal axis from the origin to the insertion and along three transversal axes from superficial to deep were found dependent on the muscle investigated. For example, the fibres of extensor digitorum longus muscle showed increased contractile and glycolytic capacities near insertion and the fibres of soleus muscle increased oxidative capacity in its middle part. Furthermore, the contribution of the fibre type that is dominant in a muscle (fast-glycolytic fibre type in extensor digitorum longus and slow-oxidative fibre type in soleus muscle) to the total number of fibres increased from origin to insertion by 15 and 30%, respectively. Along the superficial-deep axes the oxidative capacity of all fibres increased, the most in fast fibres of the soleus muscle by approximately 50%. In soleus muscle, a decrease of cross areas of all fibre types from superficial to deep was found, correlating negatively with the succinate dehydrogenase activity of the fibres. In extensor digitorum longus muscle the change in cross areas of slow-oxidative and fast-oxidative glycolytic fibres was dependent on the position of the transversal axis in the muscle. The results suggest that distribution patterns of fibre types and the metabolic make up of individual muscle fibres are adapted on the basis of local functional demands. In both muscles, higher numbers and increased oxidative capacity of fast-glycolytic fibres were found during ageing, but variations from superficial to deeper regions were irrespective of age.     

Motor denervation induces altered muscle fibre type densities and atrophy in a rat model of neuropathic pain

Loose ligation of a sciatic nerve in rats (chronic constriction injury; CCI) provokes sensory, autonomic, and motor disturbances like those observed in humans with partial peripheral nerve injury. So far, it is unknown whether these motor disturbances result from (mechanical) allodynia or from damage to the motor neuron. These considerations prompted us to assess, in CCI rats, the density of motor axons in both the ligated sciatic nerve and the ipsilateral femoral nerve. To this end, we determined the number of cholinesterase positive fibres. It has been demonstrated previously that muscle fibre type density may be used as a measure of motor denervation and/or hypokinesia. Therefore, the myofibrillar ATPase reaction was employed to assess fibre type density in biopsies obtained from the lateral gastrocnemius muscle (innervated by sciatic nerve) and rectus femoris muscle (innervated by femoral nerve). We observed axonal degeneration of motor fibres within the loosely ligated sciatic nerve, both at an intermediate (day 21) and at a late stage (day 90) after nerve injury. The reduction in the number of motor nerve fibres was more pronounced distal to the site of the ligatures than proximal. A (less pronounced) reduction of motor fibres was observed in the ipsilateral (non-ligated) femoral nerve. In line with these findings, we observed altered fibre type densities in muscle tissue innervated by the ligated sciatic nerve as well as the non-ligated femoral nerve indicative of motor denervation rather than hypokinesia. The findings of this study suggest that the motor disorder induced by partial nerve injury involves degeneration of motor nerve fibres not only within the primarily affected nerve but also within adjacent large peripheral nerves. This spread outside the territory of the primarily affected nerve suggests degeneration of motor neurons at the level of the central nervous system.     

Chromosome structure and composition

Previous studies in rodents showed a severe deterioration of pineal physiology with aging. The present study investigated the age-related changes in the content of monoamines and metabolites in rat and Syrian hamster pineal gland. In addition to melatonin, the levels of 5-hydroxytryptophan (5HTP), serotonin (5HT), 5-hydroxyindoleacetic acid (5HIAA), N-acetylserotonin (N-Ac-SHT), dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC), and noradrenaline (NA) were measured by HPLC. Pronounced reductions were found in 5HT and 5HIAA contents during daytime in rats of 24 months, which had not been observed in animals of 12 months. In addition, nighttime pineal 5HIAA, N-Ac-5HT, and melatonin contents were decreased in the old rats, although a significant day:night variation persisted. Also a diurnal fluctuation in NA, DA, and DOPAC contents was present in young and middle-aged rats but not for NA and DOPAC in the oldest rats due to a decrease in the nighttime levels. Pineal DA levels were also reduced in 24-month-old rats during the night, although a marked day:night change was still found. In the Syrian hamster pineal, significant reductions in daytime 5HT and 5HIAA were found respectively at 12 and 18 months, while nighttime levels of these compounds were decreased from 18 months. The nocturnal content of N-Ac-5HT dropped gradually from 12 months, and melatonin was reduced by 74% and 86% in hamsters of 18 and 24 months, respectively. In all these compounds, a significant day:night variation was observed irrespective of age. However, neither a day:night variation nor an effect of aging was found in terms of pineal NA content. In contrast, pineal DA and DOPAC levels displayed a diurnal variation in hamsters of 1.5 and 6 months, but not in animals of 12 and 18 months due a reduced nighttime content. These data suggest that the decline of pineal melatonin with age is a consequence of a deficit in the pathway of serotonin utilization. This probably is explained by a reduced N-acetyltransferase activity, which may be linked to impaired pineal catecholaminergic neurotransmission.     

Extravisceral masses in the peritoneal cavity: sonographically guided biopsies in 52 patients

To establish a possible correlation between the rate of cellular proliferation and already documented functional and morphological characteristics of the rat pineal gland during postnatal development, the bromodeoxyuridine labelling method was used to evaluate the fraction of cells at the S phase of the cell cycle in paraffin sections from 1-, 7-, 14- and 28-day-old rats. Numerical density, taken as an indirect measure of cell hypertrophy, was also evaluated. During the first week after birth the percentage of S phase-cells in the rat pineal gland sharply decreased from around 9% to 1.3%. A smaller but also significant decrease was found from the 7th to the 14th postnatal day where S phase cells were less than 0.5% of all pineal cells. A very low percentage was also seen in samples from 28-day-old rats. Numerical density, namely, the total number of cells per surface unit of pineal section, decreased from birth to the end of the first month. This decrease was also steeper from birth to the 7th postnatal day than at any other period of the study. These results support the idea that a strong expansion of the cellular population of the rat pineal gland precedes morphological and functional maturation and opens the way to further exploration of the relationship between functional and proliferative responses of the pineal gland.