Morphological changes induced by neuropeptide in vitro stimulation of the rat parotid gland

The effect of in vitro stimulation of rat parotid gland with the neuropeptides substance P, calcitonin gene-related peptide and galanin has been studied by microfilament fluorescence staining and in semithin sections, and compared to control incubations and in vitro stimulation with b­adrenergic and muscarinic agonists. Clear-cut aspects of massive granule exocytosis and cytoplasm vacuolation, indicative of protein and fluid secretion respectively, were obvious only after substance P stimulation, whereas treatment with galanin and calcitonin gene-related peptide produced little to no morphological changes. The results being in agreement with the outcome of other methodological approaches, these procedures appear reliable, may be effectively applied to the study of the functional regulation of secretory mechanisms, and may be particularly useful in human tissue analyses

The human nucleus cuneatus contains discrete territories that share neurochemical features with the relay nuclei for nociceptive information

Traditionally, the spinal dorsal column and the gracile (GN) and cuneate (CN) nuclei are believed to be involved in somatic tactile and proprioceptive perceptions. However, more recent clinical and experimental studies show that this system is also involved in the neurotransmission of visceral nociceptive stimuli (Willis et al., Proc. Natl. Acad. Sci. USA 96, 7675, 1999; Pale?ek J., Physiol. Res. 53, S125, 2004). Early studies in our laboratory (Del Fiacco et al., Brain Res. 264, 142, 1983; Neuroscience, 12, 591, 1984) showed that, at variance with that of laboratory animals, the human CN contains discrete subregions that are strongly immunoreactive to substance P, a neuropeptide classically involved in pain transmission. Here we provide further information on the chemical neuroanatomy of the human dorsal column nuclei and show that the substance P-immunoreactive subregions of the CN retain the neurochemical features of the protopathic relay nuclei. Tissue distribution of a number of neuropeptides, trophic factors and neuroplasticity-associated proteins was analyzed by immunohistochemistry in postmortem specimens of medulla oblongata from subjects aged 21 gestation weeks to 78 years, with no signs of neuropathology. Immunoreactivity to neuropeptides calcitonin gene-related peptide, leucine- and methionine-enkephalin, somatostatin, galanin, and peptide histidine-isoleucine, to trophins of the Neurotrophin and glial-derived neurotrophic factor families and related receptors, and to the neuroplasticity-associated proteins growth-associated protein-43 and polysialylated-neural cell adhesion molecule labels neuronal elements in restricted areas of the cuneate nucleus, located along its dorsal edge or embedded in the white matter of the cuneate fasciculus. Multiple immunolabelling shows that, with respect to one another, the examined substances are distributed in these regions as in the superficial layers of the spinal dorsal horn and trigeminal subnucleus caudalis. By contrast, the immunoreactivity in the GN is usually sparse and not gathered in definite subregions. The results show that, at variance with that of laboratory mammals, including primates, the human CN contains clear-cut subregions with neurochemical features reminiscent of those present in the relay nuclei for protopathic and pain perception. Moreover, the peculiar localization of the examined substances suggests that the superficial layers of those regions may constitute a “gelatinous subnucleus”. The origin as well as the functional involvement of such innervation remains to be elucidated

The rat dorsal column nuclei contain a region homologous to the human Locus K

Locus K is a newly identified region within the territory of the human nucleus cuneatus that shares neurochemical and histological features with protopathic second order sensory nuclei (Del Fiacco et al., 2013; Serra et al., 2013; SIAI 2014). This work is aimed at examining the rat dorsal column nuclei in order to ascertain whether a structure homologous to the human Locus K occurs in the rat brain. Rat brainstem sections were observed by means of ABC and fluorescence immunohistochemistry for neuropeptides calcitonin gene-related peptide (CGRP) and substance P (SP), Kluver-Barrera and Nissl staining. Results of the peptide immunoreactive structures in the rat dorsal column nuclei are in general good agreement with findings in previous studies (Hoeflinger et al., 1993). However, at caudalmost levels of the complex, in the territory of the cuneate fascicle and dorsal to the caudal pole of the cuneate nucleus, a small column of gray matter area can be identified that contains a dense plexus of varicose labelled nerve fibres. The observed discrete region has never distinctly described beforehand. Both its position and aspect at neuropeptide-immunoreactivity resemble those of the Locus K we detected in the human dorsal column nuclei, allowing the possibility that it represents the its homologous nucleus in the rat brain. Work funded by Fondazione Banco di Sardegna

Effect of acute administration of Pistacia lentiscus L. essential oil on rat cerebral cortex following transient bilateral common carotid artery occlusion

<p>Abstract</p> <p>Background</p> <p>Ischemia/reperfusion leads to inflammation and oxidative stress which damages membrane highly polyunsaturated fatty acids (HPUFAs) and eventually induces neuronal death. This study evaluates the effect of the administration of <it>Pistacia lentiscus </it>L. essential oil (E.O.), a mixture of terpenes and sesquiterpenes, on modifications of fatty acid profile and endocannabinoid (eCB) congener concentrations induced by transient bilateral common carotid artery occlusion (BCCAO) in the rat frontal cortex and plasma.</p> <p>Methods</p> <p>Adult Wistar rats underwent BCCAO for 20 min followed by 30 min reperfusion (BCCAO/R). 6 hours before surgery, rats, randomly assigned to four groups, were gavaged either with E.O. (200 mg/0.45 ml of sunflower oil as vehicle) or with the vehicle alone.</p> <p>Results</p> <p>BCCAO/R triggered in frontal cortex a decrease of docosahexaenoic acid (DHA), the membrane highly polyunsaturated fatty acid most susceptible to oxidation. Pre-treatment with E.O. prevented this change and led further to decreased levels of the enzyme cyclooxygenase-2 (COX-2), as assessed by Western Blot. In plasma, only after BCCAO/R, E.O. administration increased both the ratio of DHA-to-its precursor, eicosapentaenoic acid (EPA), and levels of palmytoylethanolamide (PEA) and oleoylethanolamide (OEA).</p> <p>Conclusions</p> <p>Acute treatment with E.O. before BCCAO/R elicits changes both in the frontal cortex, where the BCCAO/R-induced decrease of DHA is apparently prevented and COX-2 expression decreases, and in plasma, where PEA and OEA levels and DHA biosynthesis increase. It is suggested that the increase of PEA and OEA plasma levels may induce DHA biosynthesis via peroxisome proliferator-activated receptor (PPAR) alpha activation, protecting brain tissue from ischemia/reperfusion injury.</p

Transient receptor potential vanilloid type 1 (TRPV1) and neuropeptides in the dorsal root ganglia and spinal cord in a rat model of Bortezomib-induced neuropathy

Bortezomib (BTZ) is an effective antineoplastic drug that acts by inhibiting the ubiquitin-proteasome cellular pathways. In clinical practice, its chronic administration triggers a significant neurotoxicity, which has been associated with impairment of Aβ, Aδ, and C type primary afferent fibers, though the mechanism underlying its harmful effects remains still to be fully clarified. In order to mimic the clinical use of the drug, we have recently designed an experimental model based on the use of 0,20 mg/kg drug concentration for 8 weeks followed by a follow-up period of 4 weeks. We have previously shown that, in these conditions, a hallmark of neurotoxicity is represented by a small fiber neuropathy, whereas dorsal root ganglia (DRG) neurons did not show any morphological alterations. In order to provide data regarding the mechanism underlying BTZ harmful effects, here we characterize the spinal primary sensory neurons on the basis of their expression of the transient receptor potential vanilloid type-1 (TRPV1) and sensory neuropeptides calcitonin gene-related peptide (CGRP) and substance P (SP). In fact, TRPV1 is expressed by sensory neurons where it functions as a molecular integrator for nociception. Its activation causes depolarisation leading to burning pain and release of CGRP and SP which, in turn, activate their effector cell receptors and enhance the sensitization of nociceptors. With this aim, lumbar DRG and spinal cord of BTZ-treated model rats were processed for avidine-biotin-peroxidase complex or fluorescence immunohistochemistry. In the DRG, the immunolabelling for TRPV1 revealed a subpopulation of predominantly small- to medium-sized neurons which appeared more extensive in BTZ-treated rats. Centrally, TRPV1-LI labelled fiber tracts and terminal-like elements distributed in laminae I and II of the dorsal horn where they appeared widely codistributed with both CGRP-LI and SP-LI. With the exception of a slight more intense TRPV1 staining in lamina I of the dorsal horn of BTZ-treated vs control rats, no clear-cut differences in the distribution and amount of immunoreactivity for the three markers could be observed

Dietary essential oil components in the prevention of hypoperfusion/reperfusion-induced tissue damage in the rat cerebral cortex

To extend our previous observations on the beneficial effect of dietary Pistacia lentiscus L. essential oil during cerebral bilateral common carotid artery occlusioninduced injury, we evaluated the activity of one of its major components, beta-caryophyllene (BCP), already known to possess peculiar biological activities, in Wistar rat cerebral cortex. Cerebral hypoperfusion was produced by a 30 min bilateral common carotid artery occlusion followed by 60 min reperfusion (BCCAO/R). Animals were starved for 12 hours before surgery and, 6 hours prior to hypoperfusion, BCP (40 mg/kg/0, 45 ml of sunflower oil as vehicle) was administered via gavage. Biological samples of brain tissue, plasma and cerebrospinal fluid (CSF) were examined by HPLC, western blot, gel zymography and immunohistochemistry and analyzed for fatty acids, expression of the enzyme ciclooxygenase-2 (COX-2), CB receptors for endocannabinoids (eCBs), and peroxisome proliferator-activated receptor (PPAR)-alpha and enzymatic activity of matrix-metalloprotease-9 (MMP9). Data obtained indicate that BCP appears to influence the outcome of BCCAO/R cerebral injury by modulating changes in levels of polyunsaturated fatty acids, biosynthesis of eCBs and eCB congeners, expression of CB1 and CB2 receptors, COX-2 protein levels and enzymatic activity of MMP9. Brain tissue response to the hypoperfusion/reperfusion-induced cerebral insult is modulated by dietary administration of BCP, suggesting the possible use of this molecule as nutritional treatment in neuroprevention. Work funded by Fondazione Banco di Sardegna

Effect of acute administration of dietary Pistacia lentiscus L. essential oil on the ischemia-reperfusion-induced changes in rat frontal cortex and plasma

In this study Pistacia lentiscus L. essential oil (E.O.), a mixture of terpenes and sesquiterpenes, was tested for its protective effects in cerebral ischemia/reperfusion-induced injury in Wistar rat frontal cortex and plasma. Cerebral ischemia was produced by a 20 min bilateral common carotid artery occlusion followed by 30 min reperfusion. Pistacia lentiscus L. essential oil (E.O.) (200 mg/0, 45 ml of sunflower oil as vehicle) was administered via gavage 6 hours prior to ischemia. Rats were randomly assigned to four groups, ischemic/reperfused (I/R) and sham-operated rats treated with the vehicle or with E.O.. Different brain areas were analysed for fatty acid changes and expression of the enzyme cyclooxygenase-2 (COX-2). Ischemia/reperfusion triggered in frontal cortex a decrease of docosahexaenoic acid (DHA), the membrane highly polyunsaturated fatty acid (HPUFA) most susceptible to oxidation. Pre-treatment with E.O. prevented this change and led further to decreased levels of COX-2, as assessed by Western Blot. In plasma of ischemic/reperfused rats, E.O. administration increased both the DHA-to-eicosapentaenoic acid (EPA) ratio and levels of the endocannabinoid congeners palmytoylethanolamide (PEA) and oleoylethanolamide (OEA). The results obtained suggest that ischemia/reperfusion triggers a cerebral insult sufficient to cause a a region specific lipid peroxidation as evidenced by the detectable, significant decrease in the tissue level of DHA, the most abundant essential fatty acid of neuronal membrane phospholipids. Acute dietary pre-treatment with E.O. triggers modifications both in the frontal cortex, where COX-2 expression decreases and the decrease of DHA is apparently prevented, and in plasma, where PEA and OEA levels increase. We suggest that the activity of PEA and OEA, as endogenous ligands of the peroxisome proliferator-activated receptor (PPAR)-alpha, by inducing the peroxisomal beta oxidation, may explain the observed increase in the DHA/EPA ratio. The latter, in fact, might account for an increased metabolism of n-3 aimed at restoring DHA within damaged brain tissue. The possibility that changes in fatty acid metabolism and plasmatic availability of PEA and OEA are correlated events represents an issue worth future investigations

The distribution of proenkephalin-derived peptides in the central nervous system of turtles

The present study was carried out to examine if peptides similar to the various opioid peptide products of mammalian proenkephalin are present in the turtle central nervous system and to determine their distribution. Antisera against several enkephalin peptides were used: (1) leucine-enkephalin (LENK), (2) methionine-enkephalin (MENK), (3) methionine-enkephalin-arg 6 -phe 7 (MERF), (4) methionine-enkephalin-arg 6 -gly 7 -leu 8 (MERGL), (5) Peptide E (PEPE), and (6) BAM22P. Their specificity and cross-reactivity were carefully examined. The results indicated that LENK, MENK, and MERF (or highly similar peptides) are present in the turtle central nervous system, and that a peptide showing immunological similarity to BAM22P and PEPE also appeared to be present. In contrast, MERGL did not appear to be present. The distributions of the immunoreactive labeling for LENK, MENK, MERF, BAM22P, and PEPE were indistinguishable, and double-label studies showed that LENK, MERF, and BAM22P were colocalized within individual neurons and fibers. Although all of the above substances were observed in the same cell groups, there was some regional variation, in terms of which enkephalin peptide appeared to be most abundant. The distributions of these enkephalin peptides were very similar to those previously described in mammals and birds. Enkephalin was more abundant in the basal ganglia than in overlying telencephalic regions. Within the basal ganglia, enkephalin was present in striatal neurons and fibers and in pallidal fibers, thereby suggesting the existence of an enkephalinergic striatopallidal projection. Sensory relay nuclei of the thalamus were generally poor in enkephalinergic fibers, whereas the hypothalamus was rich in enkephalinergic neurons and fibers. Enkephalinergic neurons and fibers were present in the midbrain central gray. As is true of neurons of the nucleus spiriformis lateralis of the avian pretectum, the neurons of the homologous cell group in turtles, the dorsal nucleus of the posterior commissure of the pretectum, were found to contain enkephalin and have an enkephalinergic projection to the deep layers of the ipsilateral tectum. Enkephalinergic neurons and fibers were also abundant in the entry zones of the trigeminal nerve and dorsal root fibers of the spinal cord. The present results indicate that: (1) consistent with previously published biochemical studies (Lindberg and White, '86), proenkephalin in reptiles is similar in structure to that of mammals and, with the exception of MERGL, gives rise to similar or identical enkephalin peptides, and (2) the enkephalin peptides are found in many of the same systems of reptilian brain as mammalian and avian brain, and, therefore, may play a role in similar functions (e.g., basal ganglia motor functions) as in mammals and birds.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/50034/1/902590106_ftp.pd