Endogenous Cholinergic System Involved in Peripheral Analgesic Control in Mice Is Activated by TNF-α, CXCL-1, and IL-1ß.

INTRODUCTION: Tissue injury results in the release of inflammatory mediators, including a cascade of algogenic substances, which contribute to the development of hyperalgesia. During this process, endogenous analgesic substances are peripherally released to counterbalance hyperalgesia. The present study aimed to investigate whether inflammatory mediators TNF-α, IL-1ß, CXCL1, norepinephrine (NE), and prostaglandin E2 (PGE2) may be involved in the deflagration of peripheral endogenous modulation of inflammatory pain by activation of the cholinergic system. METHODS: Male Swiss mice were subjected to paw withdrawal test. All the substances were injected via the intraplantar route. RESULTS: The main findings of this study were as follows: (1) carrageenan (Cg), TNF-α, CXCL-1, IL1-ß, NE, and PGE2 induced hyperalgesia; (2) the acetylcholinesterase enzyme inhibitor, neostigmine, reversed the hyperalgesia observed after Cg, TNF-α, CXCL-1, and IL1-ß injection; (3) the non-selective muscarinic receptor antagonist, atropine, and the selective muscarinic type 1 receptor (m1AChr) antagonist, telenzepine, potentiated the hyperalgesia induced by Cg and CXCL-1; (4) mecamylamine, a non-selective nicotinic receptor antagonist, potentiated the hyperalgesia induced by Cg, TNF-α, CXCL-1, and IL1-ß; (5) Cg, CXCL-1, and PGE2 increased the expression of the m1AChr and nicotinic receptor subunit α4protein. CONCLUSION: These results suggest that the cholinergic system may modulate the inflammatory pain induced by Cg, PGE2, TNF-α, CXCL-1, and IL1-ß.

Altered expression of the vitamin D metabolizing enzymes CYP27B1 and CYP24A1 under the context of prostate aging and pathologies.

Low circulating levels of vitamin D are common at older ages and have been linked to an increased risk of prostate disease, including cancer. However, it has not yet been determined whether aging affects the ability of prostate cells to locally metabolize vitamin D into its active metabolite calcitriol and thus mediate the vitamin D signaling in autocrine and paracrine ways. By using a suitable rat model to interrogate spontaneous prostatic modifications over the course of aging, here we showed that both CYP27B1 and CYP24A1 enzymes, which are key players respectively involved with calcitriol synthesis and deactivation, were highly expressed in the prostate epithelium. Furthermore, as the animals aged, a drastic reduction of CYP27B1 levels was detected in total protein extracts and especially in epithelial areas of lesions, including tumors. On the other hand, CYP24A1 expression significantly increased with aging and remained elevated even in altered epithelia. Such intricate unbalance in regard to vitamin D metabolizing enzymes was strongly associated with reduced bioavailability of calcitriol in the senile prostate, which in addition to decreased expression of the vitamin D receptor, further limits the protective actions mediated by vitamin D signaling. This evidence was corroborated by the increased proliferative activity exactly at sites of lesions where the factors implicated with calcitriol synthesis and responsiveness had its expression inhibited. Taken together, our results emphasize a set of modifications over the course of aging with a high potential to hamper vitamin D signaling on the prostate. These findings highlight a crosstalk between vitamin D, aging, and prostate carcinogenesis, offering new potential targets in the prevention of malignancies and other aging-related disorders arising in the gland.

Thiamine Deficiency Increases Intrinsic Excitability of Mouse Cerebellar Purkinje Cells.

Thiamine deficiency is associated with cerebellar dysfunction; however, the consequences of thiamine deficiency on the electrophysiological properties of cerebellar Purkinje cells are poorly understood. Here, we evaluated these parameters in brain slices containing cerebellar vermis. Adult mice were maintained for 12-13 days on a thiamine-free diet coupled with daily injections of pyrithiamine, an inhibitor of thiamine phosphorylation. Morphological analysis revealed a 20% reduction in Purkinje cell and nuclear volume in thiamine-deficient animals compared to feeding-matched controls, with no reduction in cell count. Under whole-cell current clamp, thiamine-deficient Purkinje cells required significantly less current injection to fire an action potential. This reduction in rheobase was not due to a change in voltage threshold. Rather, thiamine-deficient neurons presented significantly higher input resistance specifically in the voltage range just below threshold, which increases their sensitivity to current at these critical membrane potentials. In addition, thiamine deficiency caused a significant decrease in the amplitude of the action potential afterhyperpolarization, broadened the action potential, and decreased the current threshold for depolarization block. When thiamine-deficient animals were allowed to recover for 1 week on a normal diet, rheobase, threshold, action potential half-width, and depolarization block threshold were no longer different from controls. We conclude that thiamine deficiency causes significant but reversible changes to the electrophysiology properties of Purkinje cells prior to pathological morphological alterations or cell loss. Thus, the data obtained in the present study indicate that increased excitability of Purkinje cells may represent a leading indicator of cerebellar dysfunction caused by lack of thiamine.

Características de carcaça, órgão vomeronasal, concentrações séricas de testosterona e IGF-I em machos Nelore com ductos incisivos bloqueados / Characteristics of carcass, vomeronasal organ, serum testosterone andIGF-I concentrations in Nellore males with blocked incisor ducts

Objetivou-se avaliar características quali - quantitativas da carcaça em machos Nelore, submetidos ao bloqueio dos ductos incisivos no período pré puberal. Além disso, objetivou-se avaliar as concentrações séricas de testosterona e do hormônio do crescimento semelhante a insulina do tipo I (IGF-I), e a arquitetura histológica do órgão vomeronasal (OVN). Trinta e quatro machos, no período pré puberal, foram divididos em três grupos experimentais: inteiros – animais controle sem qualquer procedimento (n = 11); bloqueados - animais com os ductos incisivos bilateralmente obstruídos (n = 10); castrados - animais com orquiepididectomia bilateral (n = 13). O OVN foi obtido no abate, ao final do experimento, para avaliação histomorfométrica. As coletas de sangue foram realizadas a cada três meses, em dois turnos (manhã e tarde), totalizando cinco avaliações. Observaram-se maiores (P0,05). Registrou-se redução da altura do epitélio neuro sensitivo do OVN nos bloqueados e castrados (P<0,05). Foi registrada menor concentração de testosterona nos bloqueados nas coletas quatro e cinco à tarde (P<0,05). Conclui-se que a obstrução dos ductos incisivos reduziu os estímulos sensoriais para o OVN, que por sua vez, promoveu alteração na concentração sérica de testosterona, sem alterar o ganho de peso dos animais, porém, não promoveu melhoria na qualidade da carcaça nas condições deste estudo.

A total of 34 prepuberal Nellore males were divided into three groups: control – animals without any procedure (n = 11); blocked - with bilaterally incisors ducts obstructed (n = 10); castrated - with bilateral orchiectomy (n = 13). This study aimed to evaluate the effect of blocking the vomeronasal organ (VNO) in qualitative and quantitative characteristics of the carcass. Also, this study aimed to evaluate testosterone and insulin-like growth factor (IGF-I) serum concentration, and the histological architecture of the OVN. Blood samples were taken every three months during two daily collections (morning and afternoon), totalizing five evaluations. It was observed higher (P 0.05). It was observed a reduction (P<0.05) of the VNO sensory epithelium height in blocked and castrated groups compared with control group. It was registered lower (P<0.05) serum testosterone concentration in blocked group at the fourth and fifth blood collection (afternoon). It was concluded that the obstruction of the incisive ducts reduced the sensorial stimuli for the OVN, which, in turn, promoted a change in the serum concentration of testosterone, without altering the weight gain of the animals, but did not promote improvement in the quality of the carcass under the conditions of this study.

Características de carcaça, órgão vomeronasal, concentrações séricas de testosterona e IGF-I em machos Nelore com ductos incisivos bloqueados / Characteristics of carcass, vomeronasal organ, serum testosterone and IGF-I concentrations in Nellore males with blocked incisor ducts

Objetivou-se avaliar características quali - quantitativas da carcaça em machos Nelore, submetidos ao bloqueio dos ductos incisivos no período pré puberal. Além disso, objetivou-se avaliar as concentrações séricas de testosterona e do hormônio do crescimento semelhante a insulina do tipo I (IGF-I), e a arquitetura histológica do órgão vomeronasal (OVN). Trinta e quatro machos, no período pré puberal, foram divididos em três grupos experimentais: inteiros ­ animais controle sem qualquer procedimento (n = 11); bloqueados - animais com os ductos incisivos bilateralmente obstruídos (n = 10); castrados - animais com orquiepididectomia bilateral (n = 13). O OVN foi obtido no abate, ao final do experimento, para avaliação histomorfométrica. As coletas de sangue foram realizadas a cada três meses, em dois turnos (manhã e tarde), totalizando cinco avaliações. Observaram-se maiores (P<0,05) pesos corporais finais (inteiros = 494,1 ± 28,71; bloqueados = 500,6 ± 23,6 e castrados = 468,3 ± 21,8 Kg) nos inteiros e bloqueados. O acabamento da carcaça foi maior nos castrados (P<0,05) em relação aos inteiros e bloqueados, enquanto o rendimento de carcaça não apresentou diferenças entre os três tratamentos (P>0,05). Registrou-se redução da altura do epitélio neuro sensitivo do OVN nos bloqueados e castrados (P<0,05). Foi registrada menor concentração de testosterona nos bloqueados nas coletas quatro e cinco à tarde (P<0,05). Conclui-se que a obstrução dos ductos incisivos reduziu os estímulos sensoriais para o OVN, que por sua vez, promoveu alteração na concentração sérica de testosterona, sem alterar o ganho de peso dos animais, porém, não promoveu melhoria na qualidade da carcaça nas condições deste estudo.

A total of 34 prepuberal Nellore males were divided into three groups: control ­ animals without any procedure (n = 11); blocked - with bilaterally incisors ducts obstructed (n = 10); castrated - with bilateral orchiectomy (n = 13). This study aimed to evaluate the effect of blocking the vomeronasal organ (VNO) in qualitative and quantitative characteristics of the carcass. Also, this study aimed to evaluate testosterone and insulin-like growth factor (IGF-I) serum concentration, and the histological architecture of the OVN. Blood samples were taken every three months during two daily collections (morning and afternoon), totalizing five evaluations. It was observed higher (P<0.05) final body weight (control= 494.1 ± 28.71; blocked = 500.6 ± 23.6; castrated = 468.3 ± 21.8 kg) at the control and blocked groups. Carcass finishing was higher in castrated animals (P <0.05), while carcass yield did not differ between treatments (P> 0.05). It was observed a reduction (P<0.05) of the VNO sensory epithelium height in blocked and castrated groups compared with control group. It was registered lower (P<0.05) serum testosterone concentration in blocked group at the fourth and fifth blood collection (afternoon). It was concluded that the obstruction of the incisive ducts reduced the sensorial stimuli for the OVN, which, in turn, promoted a change in the serum concentration of testosterone, without altering the weight gain of the animals, but did not promote improvement in the quality of the carcass under the conditions of this study.

Atrazine affects the morphophysiology, tissue homeostasis and aromatase expression in the efferent ductules of adult rats with mild alterations in the ventral prostate.

The widely used herbicide atrazine is a potent endocrine disruptor known to cause increased aromatase expression and transient increase in testicular weight followed by remarkable testis atrophy. However, whether the effects of atrazine on the testes are primary or secondary to dysfunctions in other components of male reproductive tract remains unknown. Given the high sensitivity of the efferent ductules to estrogen imbalance and the similarity to alterations previously described for other disruptors of these ductules function, and the testicular alterations observed after atrazine exposure, we hypothesized that the efferent ductules could be a target for atrazine. Herein we characterized the efferent ductules and the ventral prostate of adult Wistar rats treated with 200 mg/kg/day of atrazine for 7, 15, and 40 days. Additionally, we evaluated if the effects of atrazine in these organs could be reduced after discontinuation of the treatment. Atrazine exposure resulted in mild effects on the ventral prostate, but remarkable alterations on the efferent ductules, including luminal dilation, reduced epithelial height, and disruption of the epithelial homeostasis, which coincides with increased aromatase expression. Together with our previous data, these results suggest that at least part of the testicular effects of atrazine may be secondary to the alterations in the efferent ductules.

Persistent testicular structural and functional alterations after exposure of adult rats to atrazine.

Atrazine is an endocrine disruptor affecting testicular steroidogenesis, and promoting testicular atrophy and 3ß-HSD reduction. However, it remains unknown whether these effects are reversible or permanent. To address this issue was the aim of this study. Exposition of rats to 200mg/kg of atrazine resulted in transient increase in testicular weight, seminiferous tubules dilation and atrophy, and reduction in Leydig cell 3ß-HSD. Testicular atrophy and 3ß-HSD reduction were more pronounced after the recovery period of 75days. There was increase in aromatase expression after long-term exposure but it returned to control level after recovery. Moreover, there was increase in ED1-/ED2+, ED1+/ED2+ and ED1+/ED2- macrophages, in the recovery group. These macrophages were positive for 3ß-HSD, thereby raising possibility of their involvement in steroidogenesis. These findings further emphasize the adverse effects of atrazine on male reproduction, highlighting that testicular damages may be irreversible even after a recovery period longer than the spermatogenic cycle.

Petasiger dietz, 1909 (Trematoda: Echinostomatidae) in birds and mollusks from Brazil.

Species of Petasiger Dietz, 1909 are intestinal trematodes of aquatic birds. Despite the diversity of described species in Europe and North America, only two South American species are known and no species have been previously reported in birds from Brazil. During a study of helminths recovered from the Least Grebe, Tachybaptus dominicus (Linnaeus, 1766), from the state of Minas Gerais, Brazil, adult trematodes identified as Petasiger novemdecim Lutz, 1928 were found. The prevalence of infection was 55% (5/9), and the mean intensity of infection was 8 (1-32) parasites. Additionally, Biomphalaria straminea (Dunker, 1848) and Biomphalaria tenagophila (d'Orbigny, 1835) obtained from aquatic collections in the same state were found to harbor two species of echinostome cercariae of the Magnacauda group. These cercariae developed into metacercariae in experimentally infected Poecilia reticulata Peters, 1859. Attempts to obtain adult parasites in vertebrate host models (mice, chickens, ducks and canaries) were unsuccessful. The present study is the first report of Petasiger in Brazilian birds and the first morphological description of larvae of these parasites in Brazilian mollusks. Taxonomic aspects related to South American species of Petasiger are discussed. It is suggested that, based on significant morphological differences observed in larval stages (i.e., larval dimensions) and adult parasites (i.e., disposition of the testes), Petasiger caribbensis Nassi, 1980 should be reconsidered as a distinct species from P. novemdecim.

Ultrastructural changes in bacterial membranes induced by nano-assemblies ß-cyclodextrin chlorhexidine: SEM, AFM, and TEM evaluation.

Chemical hosts bind their guests by the same physical mechanisms as biomolecules and often display similarly subtle structure activity relationships. The cyclodextrins have found increasing application as inert, nontoxic carriers of active compounds in drug formulations. The present study was conducted to prepare inclusion complexes of chlorhexidine:ß-cyclodextrin (Cx:ß-cd), and evaluate their interactions with bacterial membrane through: scanning electron microscopy (SEM) and transmission electron microscopy (TEM); and measuring morphology alterations, roughness values, and cell weights by atomic force microscopy (AFM). It was found that the antimicrobial activity was significantly enhanced by cyclodextrin encapsulation. SEM analysis images demonstrated recognizable cell membrane structural changes and ultrastructural membrane swelling. By TEM, cellular alterations such as vacuolization, cellular leakage, and membrane defects were observed; these effects were enhanced at 1:3 and 1:4 Cx:ß-cd. In addition, AFM analysis at these ratios showed substantially more membrane disruption and large aggregates mixing with microorganism remains. In conclusion, nanoaggregates formed by cyclodextrin inclusion compounds create cluster-like structures with the cell membrane, possibly due to a hydrogen rich bonding interaction system with increasing surface roughness and possibly increasing the electrostatic interaction between cationic chlorhexidine with the lipopolysaccharides of Gram negative bacteria.

Involvement of the transepithelial calcium transport disruption and the formation of epididymal stones in roosters.

Epididymal lithiasis is a dysfunction of unknown origin characterized by the formation of calcium stones into the lumen of efferent ductules of roosters. Affected animals present an imbalance in the hormonal responsive systems that regulate the expression of proteins involved in the transepithelial calcium transport, as TRPV6, CaBP-D28K, NCX1, and PMCA. Because the efferent ductules are the major site of fluid and calcium reabsorption in excurrent ducts, it was hypothesized that impairment in local calcium homeostasis would lead to lithiasis. To test this hypothesis, we addressed the expression of these proteins in the epididymal region of affected animals. The present study focused on the investigation of the occurrence, tissue distribution, and physiological impact of the transepithelial calcium transport in roosters under normal and pathological conditions. The results showed that affected roosters presented a significant increase in TRPV6 and CaBP-D28k levels, whereas NCX1 and PMCA were not changed. Such alterations were more conspicuous in the proximal efferent ductules, in which was also observed accumulation of calcium within the epithelial cells. These findings provided the first evidences for the involvement of alteration in the expression of proteins essential for calcium reabsorption as a plausible mechanism for the formation of calcium stones within efferent ductules.

Changes in testicular morphology and steroidogenesis in adult rats exposed to Atrazine.

Atrazine is an herbicide considered as a potent endocrine disruptor, causing adverse effects on both gender of mammalian and non-mammalian species. Despite the known adverse effects of Atrazine, little is known about its action on male genital system, especially in adults. We evaluated the effects of Atrazine (50 mg/kg, 200 mg/kg and 300 mg/kg) in the 3beta-hydroxysteroid dehydrogenase (3beta-HSD) expression, plasmatic and testicular estrogen and testosterone levels, androgen receptor expression and morphological changes in adult rat testes. Atrazine at doses higher than 50 mg/kg resulted in decreased body weight, increased adrenal weight and transient increase in testis weight, followed by testis atrophy. A reduction in testosterone but increase in estradiol levels was observed. We showed for the first time that testicular 3beta-HSD protein was decreased, whereas in the adrenal it was unchanged. The results suggest that 3beta-HSD inhibition may represent an alternative mechanism through which Atrazine affects the testicular androgenesis, leading to changes in spermatogenesis.

Location of the ureteral openings in the cloacas of tinamous, some ratite birds, and crocodilians: a primitive character.

Cloacas of 67 avian species, of both sexes, from various habitats and differing dietary habits, were examined macro- and microscopically to investigate possible variation in the location of the ureteral openings. Differing from most birds studied, in adult male Rhea americana and several tinamous species the ureters were found to open into the coprodeum. In these species the urodeum receives only the vas deferens or oviduct. Similarly, in crocodiles Caiman crocodilus yacare, but not in lizards Tropidurus montanus and snakes Crotalus durissus terrificus, the ureters empty into the coprodeum. This similarity between ancient birds (ratites and tinamous) and crocodiles may indicate a primitive character linking reptiles and birds. This unusual position of the ureteral orifice can represent an adaptation to facilitate urine collection into the coprodeum and large intestine. Another possibility is that this variation in ureter position is a male reproductive strategy to avoid the mixture of urine and semen in the cloaca. There were no evident correlations between the location of the ureteral openings and the birds' habitat, diet, or histology of the coprodeal mucosa. The occurrence of a phallus in eight species of birds was detected, as well as a peculiar vascularization related to the coprodeal epithelium of anseriformes. Together, these data add to the scarce information about the morphophysiology of the avian cloaca, and also contribute to clarify avian phylogenetic linkages.

Intraepithelial plasma cells in the avian copulatory organ of two tinamou species: quantitative variation during the breeding season.

A recent observation concerning the phallus of the tinamou Nothura maculosa was the presence of cells resembling plasma cells within the epithelium of the fixed tubular portion. Owing to this unusual location of plasma cells, we studied the phalli of the tinamous N. maculosa and Rhynchotus rufescens to confirm the occurrence of intraepithelial plasma cells and to evaluate the seasonal variation in these cells. Abundant plasma cells were found within the epithelium of the fixed tubular portion of the phallus but not in the evertible portion. Migration of plasma cells from the adjacent connective tissue through the basement membrane and between the epithelial cells was frequent. Some plasma cells exhibited a rough endoplasmic reticulum with variable cisternal distension, containing fine, slightly electron-dense, granular material suggestive of immunoglobulin accumulation. An expressive increase of more than 800% in the number of intraepithelial plasma cells was found during the breeding season compared to the non-breeding season. By establishing the occurrence of intraepithelial plasma cells in the phallus and their seasonal variation, the results contribute to a better understanding of the role of these cells in the mucosal immune system of the reproductive tract of male birds.

Técnica modificada de diafanizaçäo e coloraçäo diferencial de cartilagem e osso em pequenos vertebrados / A modified technique of diaphanisation and differential staining of cartilage and bone in small vertebrates

Sao apresentadas modificaçoes de técnicas usuais de coloraçao diferencial de osso e cartilagem de pequenos vertebrados, com Alizarina Red S e Alcian Blue, respectivamente, seguidos de diafanizaçao em soluçao de NaOH-glicerina. O resultado final da técnica modificada foi a intensa coloraçao de cartilagem em azul e de osso em vermelho. O tecido adiposo apresentou-se branco-leitoso e os demais tecidos moles transparentes.