The kappa-opioid receptor is primarily postsynaptic: combined immunohistochemical localization of the receptor and endogenous opioids.

Antisera were raised against a synthetic peptide corresponding to the carboxyl terminus of the kappa-opioid receptor (KOR1). Specificity of the antisera was verified by staining of COS-7 cells transfected with KOR1 and epitope-tagged KOR1 cDNAs, by recognition by the antisera of proteins on Western blots of both transfected cells and brain tissue, by the absence of staining of both brain tissue and transfected cells after preabsorption of the antisera with the cognate peptide, and on the strong correlation between the distribution of KOR1 immunoreactivity and that of earlier ligand binding and in situ hybridization studies. Results indicate that KOR1 in neurons is targeted into both the axonal and somatodendritic compartments, but the majority of immunostaining was seen in the somatodendritic compartment. In sections from rat and guinea pig brain, prominent KOR1 staining was seen in the ventral forebrain, hypothalamus, thalamus, posterior pituitary, and midbrain. While the staining pattern was similar in both species, distinct differences were also observed. The distribution of preprodynorphin and KOR1 immunoreactivity was complementary in many brain regions, suggesting that KOR1 is poised to mediate the physiological actions of dynorphin. However, the distribution of KOR1 and enkephalin immunoreactivity was complementary in some regions as well. These results suggest that the KOR1 protein is primarily, but not exclusively, deployed to postsynaptic membranes where it mediates the effects of products of preprodynorphin and possibly preproenkephalin.

Role of dorsal raphe nucleus 5-HT(1A) and 5-HT(2) receptors in tonic immobility modulation in guinea pigs

Tonic immobility (TI) is an innate defensive behavior characterized by a state of physical inactivity and diminished responsiveness to environmental stimuli. Behavioral adaptations to changes in the external and internal milieu involve complex neuronal network activity and a large number of chemical neurotransmitters. The TI response is thought to be influenced by serotonin (5-HT) activity in the central nervous system (CNS) of vertebrates, but the neuronal groups involved in the mechanisms underlying this behavior are poorly understood. Owing to its extensive afferents and efferents, the dorsal raphe nucleus (DRN) has been implicated in a great variety of physiological and behavioral functions. in the current study, we investigated the influence of serotonergic 5-HT(1A) and 5-HT(2) receptor activity within the DRN on the modulation of TI behavior in the guinea pig. Microinjection of a 5-HT(1A) receptor agonist (8-OH-DPAT, 0.01 and 0.1 mu g) decreased TI behavior, an effect blocked by pretreatment with WAY-100635 (0.033 mu g), a 5-HT(1A) antagonist. In contrast, activation of 5-HT(2) receptors within the DRN (alpha-methyl-5-HT, 0.5 mu g) increased the TI duration, and this effect could be reversed by pretreatment with an ineffective dose (0.01 mu g) of ketanserine. Since the 5-HT(1A) and 5-HT(2) agonists decreased and increased, respectively, the duration of TI, different serotonin receptor subtypes may play distinct roles in the modulation of TI in the guinea pig. (C) 2009 Elsevier B.V. All rights reserved.

Morphometric analysis of the AMPA-type neurons in the Deiters vestibular complex of the chick brain

Chicken (Gallus gallus) brains were used to investigate the typology and the immunolabel pattern for the subunits composing the AMPA-type glutamate receptors (GluR) of hindbrain neurons of the dorsal (dND) and ventral nuclei (vND) of the Deiter`s vestibular complex (CD), which is the avian correspondent of the lateral vestibular nucleus (LVN) of mammals. Our results revealed that neurons of both divisions were poor in GluR1. The vND, the GluR2/3+ and GluR4+ label presented no area or neuronal size preference, although most neurons were around 75%. The dND neurons expressing GluR2/3 are primarily around 85%, medium to large-sized 85%, and predominantly 60% located in the medial portion of the rostral pole and in the lateral portion of the caudal pole. The majority of dND neurons containing GluR4 are also around 75%, larger (70% are large and giant), exhibiting a distribution that seems to be complementary to that of GluR2/3+ neurons. This distinct arrangement indicates functional differences into and between the DC nuclei, also signaling that such variation could be attributed to the diverse nature of the subunit composition of the GluRs. Discussion addresses the morphological and functional correlation of the avian DC with the LVN of mammals in addition to the high morphological correspondence, To include these data into the modern comparative approach we propose to adopt a similar nomenclature for the avian divisions dND and vND that could be referred as dLVN and vLVN. (C) 2008 Elsevier B.V. All rights reserved.

Maternal vitamin C deficiency during pregnancy persistently impairs hippocampal neurogenesis in offspring of guinea pigs

While having the highest vitamin C (VitC) concentrations in the body, specific functions of VitC in the brain have only recently been acknowledged. We have shown that postnatal VitC deficiency in guinea pigs causes impairment of hippocampal memory function and leads to 30% less neurons. This study investigates how prenatal VitC deficiency affects postnatal hippocampal development and if any such effect can be reversed by postnatal VitC repletion. Eighty pregnant Dunkin Hartley guinea pig dams were randomized into weight stratified groups receiving High (900 mg) or Low (100 mg) VitC per kg diet. Newborn pups (n = 157) were randomized into a total of four postnatal feeding regimens: High/High (Control); High/Low (Depleted), Low/Low (Deficient); and Low/High (Repleted). Proliferation and migration of newborn cells in the dentate gyrus was assessed by BrdU labeling and hippocampal volumes were determined by stereology. Prenatal VitC deficiency resulted in a significant reduction in postnatal hippocampal volume (P<0.001) which was not reversed by postnatal repletion. There was no difference in postnatal cellular proliferation and survival rates in the hippocampus between dietary groups, however, migration of newborn cells into the granular layer of the hippocampus dentate gyrus was significantly reduced in prenatally deficient animals (P<0.01). We conclude that a prenatal VitC deficiency in guinea pigs leads to persistent impairment of postnatal hippocampal development which is not alleviated by postnatal repletion. Our findings place attention on a yet unrecognized consequence of marginal VitC deficiency during pregnancy.

Healing of critical-size cranial defects in guinea pigs using a bovine bone-derived resorbable membrane

Purpose: To investigate the healing of critical-size cranial bone defects (9-mm-diameter) in guinea pigs treated with a bovine bone-derived resorbable membrane. Materials and Methods: A sample of 42 guinea pigs was divided into test (n = 20), control (n = 20), and standard (n = 2) groups. A full-thickness trephine defect was made in the fronto-parietal bone of each animal. In the test group, the internal and external openings of the defect were each closed with a separate membrane, and the space between them was filled with blood clot and a central spacer. In the control group, the defect was filled only with the blood clot and spacer. At 1, 3, 6, and 9 months later, the calvarias (5 per period) for both the test and control groups were collected, fixed, radiographed, and histologically processed. The Standard-group animals were sacrificed immediately after surgery and used to determine the initial size of defect radiographically. The areas of defects in the radiographs were measured with image-analysis software and were compared between groups and periods by multiple regression analysis with the Bonferroni correction. Results: At 1 and 3 months, newly formed woven bone was histologically observed in both test and control groups. Radiographically, this new bone occupied an average of 32% of the defect area at 1 month and 60% at 3 months in the test group. In the control group, 21% of the defect was filled at 1 month and 39% at 3 months. However, the differences between treatments were not statistically significant (P > .05). At 6 and 9 months, a significant increase in newly formed lamellar bone was seen histologically in both groups. Radiographically, for the test group, the new bone occupied an average of 82% of the defect area at 6 months and 96% at 9 months. For the control group, new bone composed an average of 45% of the defect area at 6 months and 40% at 9 months. The differences between the test and control groups were statistically significant at 6 and 9 months (P < .05). Complete or almost complete filling of the defect was observed in several cases. Conclusion: It was concluded that the bovine bone-derived membrane is highly biocompatible and is able to promote good healing of critical-size defects in calvaria of guinea pig.

Leishmania braziliensis ssp. in the nasal mucosa of guinea pigs inoculated in the tarsi

Two lots of 20 young male guinea pigs were inoculated subcutaneously in the tarsi with 10 (elevated to fourth potency) amastigotes of Leishmania braziliensis or L. b. guyanensis to study the susceptibility of this Neotropical hystricomorph rodent the autochthonous parasites. Almost 50% of the animals showed lesions in the inoculation site and had parazitations that were infective to hamsters, as shown by inoculating homogenates of the dermal lesion, of the spleen, of the liver, and of the nasal mucosa into hamsters at 20, 40, 60 and 120 days after inoculation of the guinea pig. Smears of the above organs showed the presence of amastigotes. Parasites inoculated into the tarsi were detected early in the skin, spleen, and liver of the guinea pig host. Blood cultures made by cardiopuncture on sacrifice of the guinea pigs were uniformly negative. The nasal mucosa of nearly all animals positive in the skin or viscera was invaded early by the parasites, although with grater frequency between 60 and 120 days post-inoculation. The use of this model for the study of mucocutaneous parasitism by L. brasiliensis is discussed, together with the phenomena of parasitism at a distance from the inoculation site, the temperature of the body regions affected, and the possible genetic influence on susceptibility of the guinea pig to L. brasiliensis.

Endogenous opiate analgesia induced by tonic immobility in guinea pigs

A function of the endogenous analgesic system is to prevent recuperative behaviors generated by tissue damage, thus preventing the emission of species-specific defensive behaviors. Activation of intrinsic nociception is fundamental for the maintenance of the behavioral strategy adopted. Tonic immobility (TI) is an inborn defensive behavior characterized by a temporary state of profound and reversible motor inhibition elicited by some forms of physical restraint. We studied the effect of TI behavior on nociception produced by the formalin and hot-plate tests in guinea pigs. The induction of TI produced a significant decrease in the number of flinches (18 ± 6 and 2 ± 1 in phases 1 and 2) and lickings (6 ± 2 and 1 ± 1 in phases 1 and 2) in the formalin test when compared with control (75 ± 13 and 22 ± 6 flinches in phases 1 and 2; 28 ± 7 and 17 ± 7 lickings in phases 1 and 2). In the hot-plate test our results also showed antinociceptive effects of TI, with an increase in the index of analgesia 30 and 45 min after the induction of TI (0.67 ± 0.1 and 0.53 ± 0.13, respectively) when compared with control (-0.10 ± 0.08 at 30 min and -0.09 ± 0.09 at 45 min). These effects were reversed by pretreatment with naloxone (1 mg/kg, ip), suggesting that the hypoalgesia observed after induction of TI behavior, as evaluated by the algesimetric formalin and hot-plate tests, is due to activation of endogenous analgesic mechanisms involving opioid synapses.

Female novelty and the courtship behavior of male guinea pigs (Cavia porcellus)

In several rodent species, an increase or recovery of sexual behavior can be observed when sexually satiated males are placed in contact with a novel mate. In order to assess the influence of female novelty on the courtship behavior of guinea pigs (Cavia porcellus), four adult males were observed during four daily 15-min sessions while interacting with the same pregnant female (same-female sessions). A new female was presented during the fifth session (switched-female session). The duration of behavioral categories was obtained from videotape records using an observational software. From the first to the second session, all males decreased the time allocated to investigating (sniffing and licking), following, and mounting the female, and that response did not recover by the end of the same-female sessions. No similar decreasing tendencies were detected in the circling or rumba categories. A marked increase of investigating occurred in all males from the last same-female session (8.1, 11.9, 15.1 and 17.3 percent session time) to the switched-female one (16.4, 18.4, 37.1 and 28.9 percent session time, respectively). Increases in following and circling were recorded in three of four males, and full-blown recovery of mounting in one male. No consistent changes in the females' responses to males (following or attacking) were observed throughout testing. These results are consistent with the hypothesis that guinea pig males recognize individual females and that courtship responses may suffer a habituation/recovery process controlled by mate novelty.

Transplantation and survival of mouse inner ear progenitor/stem cells in the organ of Corti after cochleostomy of hearing-impaired guinea pigs: preliminary results

In mammals, damage to sensory receptor cells (hair cells) of the inner ear results in permanent sensorineural hearing loss. Here, we investigated whether postnatal mouse inner ear progenitor/stem cells (mIESCs) are viable after transplantation into the basal turns of neomycin-injured guinea pig cochleas. We also examined the effects of mIESC transplantation on auditory functions. Eight adult female Cavia porcellus guinea pigs (250-350g) were deafened by intratympanic neomycin delivery. After 7 days, the animals were randomly divided in two groups. The study group (n=4) received transplantation of LacZ-positive mIESCs in culture medium into the scala tympani. The control group (n=4) received culture medium only. At 2 weeks after transplantation, functional analyses were performed by auditory brainstem response measurement, and the animals were sacrificed. The presence of mIESCs was evaluated by immunohistochemistry of sections of the cochlea from the study group. Non-parametric tests were used for statistical analysis of the data. Intratympanic neomycin delivery damaged hair cells and increased auditory thresholds prior to cell transplantation. There were no significant differences between auditory brainstem thresholds before and after transplantation in individual guinea pigs. Some mIESCs were observed in all scalae of the basal turns of the injured cochleas, and a proportion of these cells expressed the hair cell marker myosin VIIa. Some transplanted mIESCs engrafted in the cochlear basilar membrane. Our study demonstrates that transplanted cells survived and engrafted in the organ of Corti after cochleostomy.

Generation, description and storage of dendritic morphology data

It is generally assumed that the variability of neuronal morphology has an important effect on both the connectivity and the activity of the nervous system, but this effect has not been thoroughly investigated. Neuroanatomical archives represent a crucial tool to explore structure–function relationships in the brain. We are developing computational tools to describe, generate, store and render large sets of three–dimensional neuronal structures in a format that is compact, quantitative, accurate and readily accessible to the neuroscientist. Single–cell neuroanatomy can be characterized quantitatively at several levels. In computer–aided neuronal tracing files, a dendritic tree is described as a series of cylinders, each represented by diameter, spatial coordinates and the connectivity to other cylinders in the tree. This ‘Cartesian’ description constitutes a completely accurate mapping of dendritic morphology but it bears little intuitive information for the neuroscientist. In contrast, a classical neuroanatomical analysis characterizes neuronal dendrites on the basis of the statistical distributions of morphological parameters, e.g. maximum branching order or bifurcation asymmetry. This description is intuitively more accessible, but it only yields information on the collective anatomy of a group of dendrites, i.e. it is not complete enough to provide a precise ‘blueprint’ of the original data. We are adopting a third, intermediate level of description, which consists of the algorithmic generation of neuronal structures within a certain morphological class based on a set of ‘fundamental’, measured parameters. This description is as intuitive as a classical neuroanatomical analysis (parameters have an intuitive interpretation), and as complete as a Cartesian file (the algorithms generate and display complete neurons). The advantages of the algorithmic description of neuronal structure are immense. If an algorithm can measure the values of a handful of parameters from an experimental database and generate virtual neurons whose anatomy is statistically indistinguishable from that of their real counterparts, a great deal of data compression and amplification can be achieved. Data compression results from the quantitative and complete description of thousands of neurons with a handful of statistical distributions of parameters. Data amplification is possible because, from a set of experimental neurons, many more virtual analogues can be generated. This approach could allow one, in principle, to create and store a neuroanatomical database containing data for an entire human brain in a personal computer. We are using two programs, L–NEURON and ARBORVITAE, to investigate systematically the potential of several different algorithms for the generation of virtual neurons. Using these programs, we have generated anatomically plausible virtual neurons for several morphological classes, including guinea pig cerebellar Purkinje cells and cat spinal cord motor neurons. These virtual neurons are stored in an online electronic archive of dendritic morphology. This process highlights the potential and the limitations of the ‘computational neuroanatomy’ strategy for neuroscience databases.

Involvement of sensory nerves and TRPV1 receptors in the rat airway inflammatory response to two environment pollutants: diesel exhaust particles (DEP) and 1,2-naphthoquinone (1,2-NQ)

The environmental chemical 1,2-naphthoquinone (1,2-NQ) is implicated in the exacerbation of airways diseases induced by exposure to diesel exhaust particles (DEP), which involves a neurogenic-mediated mechanism. Plasma extravasation in trachea, main bronchus and lung was measured as the local (125)I-bovine albumin accumulation. RT-PCR quantification of TRPV1 and tachykinin (NK(1) and NK(2)) receptor gene expression were investigated in main bronchus. Intratracheal injection of DEP (1 and 5 mg/kg) or 1,2-NQ (35 and 100 nmol/kg) caused oedema in trachea and bronchus. 1,2-NQ markedly increased the DEP-induced responses in the rat airways in an additive rather than synergistic manner. This effect that was significantly reduced by L-732,138, an NK(1) receptor antagonist, and in a lesser extent by SR48968, an NK(2) antagonist. Neonatal capsaicin treatment also markedly reduced DEP and 1,2-NQ-induced oedema. Exposure to pollutants increased the TRPV1, NK(1) and NK(2) receptors gene expression in bronchus, an effect was partially suppressed by capsaicin treatment. In conclusion, our results are consistent with the hypothesis that DEP-induced airways oedema is highly influenced by increased ambient levels of 1,2-NQ and takes place by neurogenic mechanisms involving up-regulation of TRPV1 and tachykinin receptors.

Sequential histopathology at the Rhipicephalus sanguineus tick feeding site on dogs and guinea pigs

The tick Rhipicephalus sanguineus is a very common parasite of dogs worldwide. Dogs seem unable to acquire resistance against this tick species, whereas guinea pigs demonstrate a very strong resistance following primary infestation. We studied the inflammatory reaction at the R. sanguineus tick feeding site on dogs and guinea pigs during primary and tertiary infestations at different time intervals after attachment. Biopsies were collected after 4, 24, 48 and 96 hours. Changes that were found in all experimental groups included a cone of cement around the mouthparts of the tick, epidermal hyperplasia, edema and inflammatory cell infiltration in the dermis directly underneath the tick attachment site. Dogs reacted to ticks mainly with neutrophils, particularly after repeated exposure. Mast cells and mononuclear leukocytes were also present. Guinea pigs reacted to R. sanguineus mainly with mononuclear cells, eosinophils and basophils. These cells were particularly numerous after repeated exposure to R. sanguineus. Our results suggest that basophils and eosinophils are involved in resistance of guinea pigs to R. sanguineus and that neutrophils in dogs have little effect against this tick species.

Antibody and blood leukocyte response in Rhipicephalus sanguineus (Latreille, 1806) tick-infested dogs and guinea pigs

The dog is considered to be the natural host of Rhipicephalus sanguineus and is unable to develop appreciable resistance even after repeated feedings. The guinea pig develops strong resistance after one infestation with adult ticks. Antibody (IgG) titres against tick salivary gland antigens (SGAs) and blood leukocyte numbers in dogs and guinea pigs undergoing experimental R. sanguineus tick infestations were measured to detect a possible correlation with susceptibility or resistance of hosts. Since infested dogs develop an immediate hypersensitivity reaction to R. sanguineus antigens, total and anti-R. sanguineus SGA IgE levels were also measured in this host species. IgG and IgE antibody levels were determined by an enzyme-linked immunosorbent assay (ELISA) along three consecutive infestations of both hosts. Most dogs and guinea pigs displayed low IgG levels against R. sanguineus SGAs, though marked differences in individual response were observed. Although dog's total serum IgE levels increased significantly after infestations, no change in the amount of anti-salivary gland IgE was detected. Total and differential blood cell counts were determined in dogs and guinea pigs during primary and secondary infestation. In dogs, a tertiary infestation and a subsequent higher infestation level were also evaluated. Infested dogs did not display any alteration in blood leukocyte counts throughout the experiment. Guinea pigs, on the other hand, developed a significant basophilia during primary infestation which increased further during secondary infestation. These data reveal similarities and differences in the reactions of resistant and non-resistant hosts to ticks. They contribute for the understanding of such host-parasite relationships and will hopefully aid in the development of immune control of ticks. (C) 2003 Elsevier B.V. All rights reserved.

Guinea Pigs Develop Cutaneous Basophilia after Repeated Infestations by Nymphs of the Tick Amblyomma triste

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Immunisation of dogs and guinea pigs against Rhipicephalus sanguineus ticks using gut extract

Tick-bite naive guinea pigs were inoculated three times with Rhipicephalus sanguineus gut or salivary gland extracts and saponin as adjuvant. Dogs were inoculated three times with gut extract only as this fraction induced a more efficient resistance in guinea pigs (lower tick recovery and lower engorged female weights). Freund's adjuvant and saponin were used as adjuvants for the immunisation of dogs. Freund's adjuvant was used to enhance cellular immunity. The highest level of resistance in dogs was induced by the immunisation with gut extract and Freund's adjuvant. Many female ticks from dogs immunised this way engorged fully but died prior to oviposition. Resistant guinea pigs and dogs seemed to trigger different immune mechanisms against R. sanguineus ticks as damage to parasites also differed. A major role for cellular immunity in the resistance of dogs against R. sanguineus ticks is suggested. Resistance mechanisms against R. sanguineus ticks is discussed.