Vasoactive Intestinal Polypeptide Fibers Research Articles

026 Vasoactive Intestinal Polypeptide Directly Excites Neurons of the Subparaventricular Zone

Abstract Introduction The suprachiasmatic nucleus (SCN) is responsible for generating the circadian rhythmicity in mammals. The ventral region or core of the SCN contains neurons that express the neuropeptide vasoactive intestinal polypeptide (VIP). VIP signaling is central for coherency and synchrony of SCN activity. VIP-expressing neurons in the SCN densely project to the ventral subregions of the subparaventricular zone (vSPZ). We studied the effects of VIP on vSPZ neurons in brain slices of mice with a combined calcium imaging and whole-cell patch-clamp recording techniques. We used calcium imaging to assess the effects of VIP on vSPZ neurons as a population and we acquired patch-clamp recordings to explore the effects of VIP on the electrical properties and the synaptic inputs to vSPZ neurons. Methods We expressed GCamp6 in vSPZ neurons by stereotaxically injecting AAV10-DIO-Ef1a-GCamp6 into the vSPZ of vGAT-IRES-Cre mice. Brain slices were prepared two weeks later and images were captured using a standard GFP filter set. We performed whole-cell recordings of the vSPZ neurons of wild-type mice. We assessed the effects of VIP on the membrane potential and the on excitatory synaptic input in vSPZ neurons. Results Using GCamp6-based in vitro calcium imaging we found that VIP excites 17% of vSPZ neurons and this effect was maintained in the presence of tetrodotoxin (TTX) and synaptic blockers for AMPA/NMDA and GABAA transmissions suggesting a direct effect of VIP on vSPZ neurons. We confirmed this result with patch-clamp recordings. We found that 29% of vSPZ neurons were excited by VIP. VIP produced a membrane depolarization of vSPZ neurons in the presence of antagonists for AMPA/NMDA and GABAA receptors. In addition, we found that in a small percentage of vSPZ neurons VIP increased the frequency of the glutamatergic excitatory postsynaptic currents, suggesting an additional excitatory mechanism. Conclusion Our results demonstrate that exogenous VIP directly excites the vSPZ neurons producing an increase in intracellular calcium and membrane depolarization. In addition, VIP increases glutamatergic afferent inputs to vSPZ neurons indicating an additional synergistic excitation. We conclude that when VIP is released from the SCN VIP fibers it can activate vSPZ neurons. Support (if any) NS091126 and HL149630.

Localization of Vasoactive Intestinal Polypeptide Receptor 1 (VPAC1) in Hypothalamic Neuroendocrine Oxytocin Neurons; A Potential Role in Circadian Prolactin Secretion.

Prolactin (PRL) is a versatile hormone and serves a broad variety of physiological functions besides lactation. The release of PRL from lactotrophs in the pituitary has in rodents been shown to be released with a circadian pattern depending on the physiological state of the animal. The circadian release of PRL seems to be complex involving tonic inhibition by dopamine (DA) neurons on lactotrophs and one or even several releasing factors. Because of the circadian releasing pattern of PRL, neurons in the suprachiasmatic nucleus (SCN), “the brain clock,” and especially the neurons expressing neuropeptide vasoactive intestinal polypeptide (VIP), have been suggested to be involved in the circadian regulation of PRL. In the present study, we used fluorescence immunohistochemistry, in situ hybridization histochemistry, confocal microscopy, three-dimensional reconstruction, and highly specific antibodies to visualize the occurrence of VIP receptors 1 and 2 (VPAC1 and VPAC2) in mouse brain hypothalamic sections stained in combination with VIP, oxytocin (OXT), arginine vasopressin (AVP), and DA (tyrosine hydroxylase, TH). We demonstrated that VIP fibers most likely originating from the ventral part of the SCN project to OXT neurons in the magnocellular part of the paraventricular nucleus (PVN). In the PVN, VIP fibers were found in close apposition to OXT neuron exclusively expressing the VPAC1 receptor. Furthermore, we demonstrate that neither OXT neurons nor TH or AVP neurons were expressing the VPAC2 receptor. VPAC1 receptor expression was also found on blood vessels but not in neurons expressing AVP or TH. These findings suggest that VIP signaling from the SCN does not directly target DA neurons involved in PRL secretion. Furthermore, the findings support the notion that VIP from neurons in the SCN could regulate circadian release of OXT in the posterior pituitary or modulate OXT neurons as a releasing factor involved in the circadian regulation of PRL from pituitary lactotrophs.

The Role of VIP in Social Behavior: Neural Hotspots for the Modulation of Affiliation, Aggression, and Parental Care.

Although the modulation of social behaviors by most major neurochemical systems has been explored, there are still standouts, including the study of vasoactive intestinal polypeptide (VIP). VIP is a modulator of circadian, reproductive, and seasonal rhythms and is well known for its role in reproductive behavior, as it is the main vertebrate prolactin-releasing hormone. Originally isolated as a gut peptide, VIP and its cognate receptors are present in virtually every brain area that is important for social behavior, including all nodes of the core "social behavior network" (SBN). Furthermore, VIP cells show increased transcriptional activity throughout the SBN in response to social stimuli. Using a combination of comparative and mechanistic approaches in socially diverse species of estrildid finches and emberizid sparrows, we have identified neural "hotspots" in the SBN that relate to avian affiliative behavior, as well as neural "hotspots" that may represent critical nodes underlying a trade-off between aggression and parental care. Specifically, we have found that: (1) VIP fiber densities and VIP receptor binding in specific brain sites, such as the lateral septum, medial extended amygdala, arcopallium, and medial nidopallium, correlate with species and/or seasonal differences in flocking behavior, and (2) VIP cells and fibers within the anterior hypothalamus-caudocentral septal circuit relate positively to aggression and negatively to parental care while VIP elements in the mediobasal hypothalamus relate negatively to aggression and positively to parental care. Thus, while a given behavior or social context likely activates VIP circuitry throughout the SBN and beyond, key brain sites emerge as potential "hotspots" for the modulation of affiliation, aggression, and parental care.

Substance P (SP) and vasoactive intestinal polypeptide (VIP) in the lower uterine segment in first and repeated cesarean sections

The authors studied the presence of substance P (SP) and vasoactive intestinal polypeptide (VIP) and their related fibers in the lower uterine segment (LUS) in 133 women undergoing cesarean sections (CS) during active labor. These were divided into 2 groups: women undergoing repeat or first CSs. Specimens were collected from the LUS and were evaluated by light microscopy and by immunohistochemistry, for the morphometrical quantification of the SP and VIP fibers in the LUS. The SP amount was higher in the post-CS scar, while the VIP amount decreased: nerve fibers contained an SP amount of up to 13 ± 2.6 C.U., while nerve fibers contained a VIP amount of up to 7 ± 1.9 C.U. The SP amount counts 10 ± 1.5% of the total Bodian fibers, and the ratio of the VIP is 10 ± 1.8% of their total amount. In normal conditions only 6.61 C.U. of the Bodian surface is occupied by SP related nerve fibers in contrast to 6.63 C.U. of the total surface by VIP; the amount of SP increased up to 13 ± 2.6 C.U., while it decreased in the LUS previous scars. The SP levels are higher in repeat CS, while the VIP levels are reduced in the LUS. The increase of SP is probably linked to the attempt to achieve cervical ripening in a post-CS LUS, with the possible consequences of dystocia during vaginal birth after CS. Nevertheless, the decrease of VIP probably affects the relaxation of the internal uterine orifice, compromising the LUS formation and cervical ripening.

Sodium Depletion Increases Sympathetic Neurite Outgrowth and Expression of a Novel TMEM35 Gene-Derived Protein (TUF1) in the Rat Adrenal Zona Glomerulosa

The adrenal zona glomerulosa (ZG) secretes aldosterone to regulate sodium balance. Chronic sodium restriction increases aldosterone accompanied by ZG expansion. The ZG is innervated by sympathetic, vasoactive intestinal polypeptide (VIP) and neuropeptide tyrosine (NPY), and sensory, calcitonin gene-related peptide, nerves. It is unclear whether innervation is affected by ZG growth. Therefore, we measured neurite outgrowth in the ZG of adult male rats after dietary sodium manipulation. In response to 1 wk sodium restriction, VIP and NPY fibers elongated in parallel with expansion of the ZG, shown by aldosterone synthase (AS) expression, but calcitonin gene-related peptide fibers were not affected. Sodium repletion resulted in parallel regression in VIP and NPY fiber length and AS expression. These results show that sympathetic, but not sensory, innervation is coordinated with ZG growth. Mediators underlying changes in innervation are unknown; therefore, we characterized a novel gene TMEM35 [termed the unknown factor-1 (TUF1) due to its unknown function] that shows extensive overlap with AS in ZG. After sodium restriction, TUF1 expanded in parallel with the ZG. TUF1 bound the low-affinity neurotrophin receptor, p75NTR, which was expressed in NPY fibers and showed a response similar to TUF1 after sodium manipulation. TUF1- p75NTR binding was competitively displaced by nerve growth factor but not by TUF1 lacking the p75NTR binding motif. Moreover, TUF1 mRNA in rat ZG cells increased after angiotensin II exposure in vitro. Collectively, these findings suggest that TMEM35/TUF1 is a candidate for modulating neurite outgrowth in the ZG after sodium depletion.

Re-innervation pattern of the ‘neovagina’ created from the bladder flap in patients with Mayer-Rokitanski-Kistner-Hauser syndrome: An immunochemical study

Background. Cystovaginoplasty (CVP) is a method of vaginal reconstruction in women with Mayer-Rokitansky-Kistner-Hauser Syndrome (MRKHS). The neo-vagina allows normal sexual intercourses, but after CVP, the sexual life of patients with MRKHS does not differ significantly from normal females. Therefore, we decided to elucidate the pattern of sensory re-innervation of the bladder flap used for the surgery.Methods. Biopsies were taken from vaginal vestibule and urinary bladder during the CVP and 1 year later in four patients with MRKHS. The following neurotransmitters were studied calcitonin gene-related peptide (CGRP), galanin (GAL), vasoactive intestinal polypeptide (VIP) and pituitary adenylate cyclase-activating peptide (PACAP).Results. CGRP and PACAP nerve fibres were sparse under the urothelium and in submucosal layer of the neovagina, they were more numerous around blood vessels and in the vicinity of smooth muscles. This was similar to the pattern observed in the urinary bladder. VIP- and GAL-positive nerve fibres were most numerous in the submucosa around blood vessels and in smooth muscle bundles of neovagina. They were distinctly less numerous beneath the epithelium. This innervation pattern mimicked that seen in normal vagina and in vaginal vestibule of patients with MRKHS.Conclusions. Our findings demonstrate considerable nervous system plasticity in the bladder flap. Distribution of presumably sensory CGRP and PACAP immunoreactive nerve fibers was similar to the pattern observed within the intact bladder wall, and VIP or GAL immunoreactive nerve fibers (vasomotor functions) were distributed in a manner similar to that observed in the intact vaginal wall.

Vasoactive intestinal polypeptide (VIP) and VPAC1 receptor in adult human dental pulp in relation to caries

To quantitatively measure VIP levels and to qualitatively study the distribution of VIP fibres and demonstrate the presence of the VPAC1 receptor in human dental pulp from carious and non-carious adult human teeth. Dental pulp samples were collected from non-carious, moderately carious and grossly carious adult human teeth. VIP levels were determined using radioimmunoassay. The distribution of VIP fibres was studied using immunohistochemistry. The VPAC1 receptor protein expression was determined by Western blotting. VIP levels were found to be significantly elevated in the dental pulp of moderately carious compared with non-carious (p=0.0032) or grossly carious teeth (p=0.0029). The distribution of VIP fibres was similar in non-carious and carious teeth, except that nerve bundles appeared thicker in the pulp samples from carious compared with non-carious teeth. Western blotting indicated that the VPAC1 receptor proteins were detected in similar levels in pooled dental pulp samples from both carious and non-carious teeth. It is concluded that quantitative changes in the levels of VIP in human dental pulp during the caries process and the expression of VPAC1 receptor proteins in membrane extracts from carious and non-carious teeth suggests a role for VIP in modulating pulpal health and disease.

Arginine vasopressin and vasoactive intestinal polypeptide fibers make appositions with gonadotropin-releasing hormone and estrogen receptor cells in the diurnal rodent Arvicanthis niloticus

Diurnal and nocturnal animals differ with respect to the timing of a host of behavioral and physiological events including those associated with estrus, but the neural bases of these differences have not been elucidated. We investigated this issue by examining the distribution of cells containing gonadotropin-releasing hormone (GnRH) as well as estrogen receptors (ERs) in relation to fibers containing peptides present in the suprachiasmatic nucleus (SCN) in a diurnal animal, Arvicanthis niloticus (the unstriped Nile grass rat). We found that fibers containing two peptides found in SCN cells, arginine vasopressin and vasoactive intestinal polypeptide appeared to be in contact with GnRH and ER positive cells. These data suggest that temporal information is carried along the same direct pathways from the SCN to GnRH and ER neurons in day- and night-active species.

Stereological Quantification of Nerve Fibers Immunoreactive to PGP 9.5, NPY, and VIP in Rat Prostate During Postnatal Development

This work was undertaken to study prostate innervation during the postnatal development of rats. It deals with the quantification of nervous fibers throughout all the regions of the rat prostate during the postnatal development using a general marker for nervous tissue, protein gene product 9.5, and 2 neuropeptides (NPY and VIP). Forty male Wistar rats (prepubertals, pubertals, young, and aged adults) were studied for immunohistochemistry of protein gene product (PGP 9.5), neuropeptide Y (NPY), and vasoactive intestinal polypeptide (VIP). They were also evaluated for length density of nerve fibers (L(V) PGP 9.5, L(V) NPY, L(V) VIP). Nerve fibers immunoreactive to the 3 antigens studied were detected in all the groups and in all the prostate zones. Periductal L(V) NPY evidenced a significant increase in the pubertal group, maintained throughout adult life. Periductal L(V) VIP showed a significant increase in young adults. The length densities of VIP and NPY fibers were significantly higher in periductal and ampular locations in comparison with dorsal and ventral sites. It can be concluded that the relative amount of nerve fibers in rat prostate, detected by PGP 9.5, does not change during postnatal development. There were significant changes in NPY and VIP fibers, showing an increase in periurethral ducts at puberty. The abundance of peptidergic innervation around the excretory ducts is related to their contractility. The development of innervation of periurethral ducts is regulated by androgens.

Peptidergic innervation in the rat carotid body after 2, 4, and 8 weeks of hypocapnic hypoxic exposure.

The distribution and abundance of neuropeptide-containing nerve fibers were examined in the carotid bodies of rats exposed to hypocapnic hypoxia (10% O2 in N2) for 2, 4, and 8 weeks. The carotid bodies after 2, 4, and 8 weeks of hypoxic exposure were enlarged by 1.2-1.5 times in the short axis, and 1.3-1.7 times in the long axis in comparison with the normoxic control ones. The enlarged carotid bodies contained a number of expanded blood vessels. Mean density per unit area (10(4) microm2) of substance P (SP) and calcitonin gene-related peptide (CGRP) immunoreactive fibers was transiently high in the carotid bodies after 4 weeks of hypoxic exposure, and decreased significantly to nearly or under 50% after 8 weeks of hypoxic exposure. Density of vasoactive intestinal polypeptide (VIP) immunoreactive fibers increased significantly in all periods of hypoxic exposure observed, and was especially high in the carotid bodies after 4 weeks of hypoxic exposure. Density of neuropeptide Y immunoreactive fibers was unchanged in the carotid bodies during hypoxic exposure. These characteristic changes in the density of SP, CGRP, and VIP fibers in the carotid bodies after 4 weeks of hypoxic exposure suggest that the role of these neuropeptide-containing fibers may be different in the carotid bodies after each of three periods of hypoxic exposure, and that the peptidergic innervation after 8 weeks of hypoxic exposure may show an acclimatizing state.

Mast cells, nerves and neuropeptides in atopic dermatitis and nummular eczema.

The association between mast cells and sensory nerves and the distribution of the neuropeptides substance P (SP), vasoactive intestinal polypeptide (VIP) and calcitonin gene-related peptide (CGRP) were studied immunohistochemically in lesional and nonlesional skin of 26 atopic dermatitis (AD) and 23 nonatopic nummular eczema (NE) patients. Mast cell-nerve contacts were counted morphometrically and confirmed by confocal laser scanning microscopy. Neuropeptide positivity was assessed semiquantitatively. Dermal contacts between mast cells and nerves were increased in number in both lesional and nonlesional samples of AD and NE when compared to those in normal controls, although only the values in lesional AD reached statistical significance ( P<0.05). Nerve-mast cell contacts in the basement membrane zone were seen practically only in lesional NE. SP and CGRP fibres were prominently increased in lesional samples when compared to their nonlesional controls both in AD and NE in the epidermis and in the papillary dermis. In both AD and NE, only small differences were found regarding VIP positivity in lesional and nonlesional biopsies. The epidermis was devoid of VIP positivity. In conclusion, SP and CGRP but not VIP fibres were more frequent in lesional than in nonlesional papillary dermis of both AD and NE. Since mast cells are also increased in number in lesions of AD and NE, they are able to maintain neurogenic inflammation through activation by SP and CGRP. The increased SP/CGRP nerves in the epidermis of AD and NE lesions may stimulate keratinocytes to release cytokines which affect various cell types enhancing inflammation.

Pituitary adenylate cyclase-activating polypeptide does not colocalize with vasoactive intestinal polypeptide in the hypothalamic magnocellular nuclei and posterior pituitary of cats and rats.

Pituitary adenylate cyclase-activating polypeptide (PACAP) and vasoactive intestinal polypeptide (VIP) immunoreactive cells were demonstrated in the hypothalamic magnocellular nuclei in cats and rats. In cats these immunoreactive cells were stained without any treatment or intervention; however, in rats we had to use the pituitary stalk section to enhance the amount of PACAP and VIP for successful immunostaining. In both species the regions occupied by PACAP and VIP immunoreactive cells partially overlap each other in the paraventricular and supraoptic nuclei. Nevertheless, in either cats or rats PACAP and VIP immunoreactivities do not colocalize in the same cells studied by double labeling immunohistochemistry (IHC) or by the combination of immunohistochemistry and in situ hybridization. As was expected, PACAP and VIP immunoreactive materials were stored in different fibers of the posterior pituitary where the distribution of PACAP and VIP fibers also showed different patterns: PACAP fibers form a dense plexus at the periphery of the posterior lobe, in the vicinity of the intermediate lobe; however, the VIP fibers were evenly distributed mainly in the center of the posterior lobe. In spite of the high sequence homology of PACAP and VIP, the two peptides are synthesized in different subpopulations of hypothalamic neurons. This different distribution correlates well with the different role of the hypothalamic PACAP and VIP in the biologic clock and in the functions of the anterior and posterior pituitary.

Fos-induction in gonadotropin-releasing hormone neurons receiving vasoactive intestinal polypeptide innervation is reduced in middle-aged female rats.

A hallmark of reproductive aging in rats is a delay in the initiation and peak, and a decrease in the amplitude, of both proestrous and steroid-induced surges of LH and a decrease in the number of GnRH neurons that express Fos during the surge. The altered timing of the LH surge and the decline in Fos expression in GnRH neurons may be due to changes in the rhythmic expression of vasoactive intestinal polypeptide (VIP), a neuropeptide that carries time-of-day information from the circadian pacemaker, located in the suprachiasmatic nuclei (SCN), to GnRH neurons. The goals of our study were to determine if aging alters 1) the innervation of GnRH neurons by VIP and 2) the ability of VIP to activate GnRH neurons by examining the effects of aging on the number of GnRH neurons apposed by VIP fibers and the number of GnRH neurons that receive VIP input that express Fos. Immunocytochemistry for GnRH and VIP; or GnRH, VIP, and Fos was performed on tissue sections collected from young (2-4 mo), regularly cycling females and middle-aged (10-12 mo) females in constant estrus. The number of GnRH neurons, GnRH neurons apposed by VIP fibers, and GnRH neurons that express Fos and apposed by VIP fibers were counted in both age groups. Our results clearly demonstrate that aging does not alter the number of GnRH neurons that receive VIP innervation. However, the number of GnRH neurons that receive VIP innervation and coexpress Fos decreases significantly. We conclude that the age-related delay in the timing of the LH surge is not due to a change in VIP innervation of GnRH neurons, but instead may result from a decreased sensitivity of GnRH neurons to VIP input.

Comparative study on the appearance of various bioactive peptides in foregut derivates during the ontogenesis

Bioactive peptides have an important multifunctional role in the gastrointestinal tract. In the present study we have investigated the dynamism of the appearance of PACAP (pituitary adenylate cyclase activating polypeptide), VIP (vasoactive intestinal polypeptide), gastrin, and secretin immunoreactivities in human foregut derivates during the ontogenesis using an immunohistochemical approach. None of these peptides were observed in the foregut derivates of an 8-week-old embryo. VIP immunoreactive nerve fibers appeared by the 11th week in the smooth muscle layers of the stomach. No other peptide immunoreactivities were observed of this stage. In 18- and 20-week old fetuses PACAP, secretin, and gastrin immunoreactive cells appeared in the developing glands of the stomach. In the duodenum gastrin immunoreactivity was present in the Lieberkühn's glands and secretin immunoreactive cells were seen between the surface epithelial cells. In the pancreas secretin immunoreactivity was found in the Langerhans islets; however, PACAP immunreactivity was observed in the exocrine portion. The distribution of VIP fibers did not change during the fetal life and it was similar to the adult pattern. According to our results the appearance of PACAP, secretin, and gastrin in the developing glands suggests their role in the proliferation and differentiation of the epithelial derivates.

VIP innervation: sharp contrast in fetal sheep and baboon adrenal glands suggests differences in developmental regulation

Immunocytochemical technique and light microscopy were used to ascertain the relationship between vasoactive intestinal polypeptide (VIP) and tyrosine hydroxylase in fetal sheep and fetal baboon adrenal cortices and medullae at 85% of gestation. VIP immunostaining was extremely robust in fetal sheep adrenal cortical neurofibers and cells while weak in fibers and nonexistent in cells of fetal baboon. Also, tyrosine hydroxylase-immunopositive cells, present throughout the adrenal cortices of both fetal sheep and baboons, were heavily innervated by VIP-immunoreactive neurofibers in fetal sheep, but not in fetal baboons. Adrenal cortical VIP-immunopositive fibers occurred in greater ( P<0.05) frequency in fetal sheep than in fetal baboons (14.82±3.10 vs. 0.84±0.26 fibers/field), were larger in diameter (2.93±0.34 vs. 0.93±0.07 μm) and ran for longer distances in the plane of section (127.85±5.16 vs. 74.53±4.93 μm). VIP immunogenicity in cells (ganglion and chromaffin) and fibers was robust in fetal adrenal medulla of sheep while nonexistent in baboons. VIP fibers in fetal sheep medulla were smaller in diameter compared to fetal sheep cortex (1.22±0.13 vs. 2.93±0.34 μm, P<0.05), but not compared to extrinsic nerve fibers (1.30±0.09 μm). We hypothesize that in fetal sheep of this age, medullary neurofibers derive primarily from extrinsic sources while cortical fibers arise from cortical ganglion cells. We conclude that at 85% of gestation the potential for VIP neural control of paracrine (e.g., glucocorticoid/catecholamine) interactions in both adrenal cortex and medulla is much greater in fetal sheep compared to fetal baboons.

Neurochemical and cellular markers in human cervix of late pregnant, postpartal and non-pregnant women

The aim was to evaluate the peptidergic innervation and the dendritic cell content in the cervix uteri. Cervical biopsies were obtained from late pregnant (n=5), postpartal (n=5) and non-pregnant (n=5) women. The samples were prepared for immunohistochemistry using antibodies to protein S-100 (S-100), calcitonin gene-related peptide (CGRP), vasoactive intestinal polypeptide (VIP), human peptide histidine isoleucine amide (PHM 27), neuropeptide tyrosine (NPY), and human histocompatibility complex class II subregion DR (HLA-DR). Nerve fibers positive for protein S-100, and dendritic cells positive for S-100 and HLA-DR were abundant in the cervix, especially at late pregnancy. CGRP, VIP, PHM-27 and NPY positive nerve fibers were present in non-pregnant, short nerve fibers and scattered immunoreactivity at term, and further scattered immunoreactivity after parturition. NPY positive nerve fibers were decreased at term, and after parturition a scattered immunoreactivity was observed. The abundant protein S-100 positive nerve fibers implies an impact of myelinated nerves in the cervix uteri during pregnancy. The abundant dendritic cells, positive for HLA-DR and S-100, especially at term, indicates a general activation of the immune system until late pregnancy and parturition. The changed occurrence and distribution of immunoreactivity for CGRP, VIP and PHM-27 suggest a release of these neuropeptides until term. The changes in NPY immunoreactivity indicate a release of NPY around parturition.

Vasoactive intestinal polypeptide in sacral primary sensory pathways in the cat.

Unmyelinated sensory axons in the sacral spinal cord may play a role in bladder reflexes under certain pathological conditions. Previous data suggested vasoactive intestinal polypeptide (VIP) might be contained exclusively in sensory C-fibers, some of which innervate the bladder. This study was undertaken to describe the morphology of these VIP fibers in the sacral cord of the cat. VIP immunoreactivity was confined to unmyelinated axons observed at several levels of the sensory pathway including the dorsal root ganglia, dorsal roots, Lissauer's tract, and the lateral collateral pathway. A combination of light and electron microscopic observations showed VIP-immunoreactive fibers with labeled varicosities and synaptic terminals in laminae I, IIo, V, VII, and X. VIP-immunolabeled varicosities had a mean diameter of 1.6 microm (range = 0.11-7.4 microm, S.D. = 1.01, n = 311) with a small percentage (8%) being relatively large (3-7.4 microm). VIP varicosities contained a mixture of small clear vesicles (CLV) and large dense core vesicles (LDV). Although most varicosities contained a moderate number of LDVs (14.86 LDVs/microm2), some varicosities contained a large number of LDVs, whereas others contained very few. Varicosities that possessed synaptic specializations were classed as terminals and were divided into three morphological classes. Two of these resembled Gray's Type I terminal, whereas a third was similar to the Gray's Type II terminal. There was no consistent relationship between vesicle content of the terminal and the type of synaptic contact it possessed. This study shows that in the sacral spinal cord of the cat, VIP terminals originate only from C-fibers, terminate primarily in laminae I and V, and exhibit a variety of morphologies consistent with heterogeneous origins and functions of the lower urinary tract.

Fetal tissue containing the suprachiasmatic nucleus restores multiple circadian rhythms in old rats.

The suprachiasmatic nucleus (SCN) is the major circadian pacemaker in mammals. When fetal tissue containing the SCN is transplanted into young rats whose circadian rhythms have been abolished by SCN lesions, the rhythms gradually reappear. Circadian rhythms in many rats deteriorate or disappear with age. The rationale of the present study was that old rats with poor circadian rhythms resemble young rats with SCN lesions. If there is a similar mechanism underlying this resemblance, then fetal tissue containing the SCN should restore rhythms in old rats. Therefore, we implanted such tissue into the third ventricle of intact aged rats with poor circadian rhythms. Body temperature, locomotor activity, and/or drinking were measured simultaneously within subjects. Grafts and hosts were stained immunocytochemically for vasoactive intestinal polypeptide (VIP), arginine vasopressin (AVP), and neuropeptide Y (NPY). Of 23 SCN grafts, 14 were viable (cells observable with Nissl or peptide staining). In 7 of the 14 aged hosts, up to three circadian rhythms were improved or restored. VIP cells were always observable, which was not the case for AVP cells or NPY fibers. In the other seven hosts, no circadian rhythm was improved. Compared with the successful grafts, these unsuccessful grafts had similar amounts of AVP and NPY staining but significantly less VIP cell and/or fiber staining. Fetal cerebellar grafts, which do not contain any of the three peptides, did not improve or restore any rhythms. Thus the degeneration of circadian rhythms in aged rats may be due, at least in part, to deterioration of the aged SCN and in particular, to a loss of function of VIP-containing neurons.

Hypoxic adaptation of the peptidergic innervation in the rat carotid body

The abundance of substance P (SP)-, calcitonin gene-related peptide (CGRP)-, vasoactive intestinal polypeptide (VIP)-, and neuropeptide Y (NPY)-immunoreactive nerve fibers in the carotid body was compared between normoxic and chronically hypoxic rats (10% O 2 and 3.0–4.0% CO 2 for 3 months). The immunoreactive fibers appeared as thin processes with many varicosities, and were distributed mainly around the vasculatures. In the normoxic control carotid body, NPY fibers were more numerous than VIP, CGRP, and SP fibers. In the chronically hypoxic rats, the carotid body was enlarged several fold, and the mean absolute number of VIP and NPY fibers was 3.88 and 2.22 times higher than in the normoxic carotid body, respectively, although that of SP and CGRP fibers was not changed. When expressed as density per unit area of the parenchyma, the density of SP and CGRP fibers in the chronically hypoxic carotid body decreased significantly to under 50%, the density of VIP fibers increased significantly 1.80 times, and the density of NPY fibers were unchanged. Immunoreactivity for four neuropeptides was not found in the glomus cells of normoxic or chronically hypoxic carotid bodies. These results suggest that altered peptidergic innervation of the chronically hypoxic carotid body is one feature of hypoxic adaptation. Because these neuropeptides are vasoactive in nature, altered carotid body circulation may contribute to modulation of the chemosensory mechanisms by chronic hypoxia.

Distribution of vasopressin and vasoactive intestinal polypeptide (VIP) fibers in the human hypothalamus with special emphasis on suprachiasmatic nucleus efferent projections.

The human suprachiasmatic nucleus (SCN) is located in the basal part of the anterior hypothalamus and is considered as the biological clock that generates circadian rhythms and synchronizes the daily activity pattern with the environmental light-dark cycle. However, the mechanisms and pathways by which the SCN transmits its information to the other brain areas are unknown. Therefore, in the present study, we investigated the efferent projections of the SCN by the immunocytochemical staining of two major peptidergic SCN neurotransmitters: vasopressin (VP) and vasoactive intestinal polypeptide (VIP). It confirmed that these peptides are present in different subdivisions of the SCN. The results of this investigation show that VP and VIP fibers arising from the SCN were detected to branch extensively and hence seem to innervate the SCN itself and the central and medial part of the anteroventral hypothalamic area (AVH), the area below the paraventricular nucleus (sub-PVN), the ventral part of the paraventricular nucleus (PVN), and the dorsomedial nucleus of the hypothalamus (DMH). There appeared to be substantial congruity between the presumptive human SCN projections and those as observed by tracing in rat or hamster. Regarding the anatomical organization of the human SCN projections, the main projection areas appeared to be the AVH, the sub-PVN, the ventral part of the PVN, and the DMH. The observation that VIP and in particular VP fibers pass between the SCN and the PVN suggests that the human SCN and the PVN may have a direct anatomical connection. In addition, VP and VIP fibers were detected in several other hypothalamic areas that are not known to have clear direct connections to the SCN. The possible origin of these VP and VIP fibers is discussed.