Post-weaning social isolation modifies neonatal anoxia-induced changes in energy metabolism and growth of rats.

Neonatal oxygen deficiency in rats may disturb growth and long-term metabolic homeostasis. In order to facilitate metabolic evaluation, the subjects are usually housed individually. However, social isolation associated with individually housed conditions alters animal behavior, which may influence the experimental results. This study investigated the effects of social isolation on neonatal anoxia-induced changes in growth and energy metabolism. Male and female Wistar rats were exposed, on postnatal day 2 (P2), to either 25-min of anoxia or control treatment. From P27 onward, part of the subjects of each group was isolated in standard cages, and the remaining subjects were housed in groups. At P34 or P95, the subjects were fasted for 18 h, refeed for 1 h, and then perfused 30 min later. Glycemia, leptin, insulin, and morphology of the pancreas were evaluated at both ages. For subjects perfused at P95, body weight and food intake were recorded up to P90, and the brain was collected for Fos and NeuN immunohistochemistry. Results showed that male rats exposed to neonatal anoxia and social isolation exhibited increased body weight gain despite the lack of changes in food intake. In addition, social isolation (1) decreased post-fasting weight loss and post-fasting food intake and (2) increased glycemia, insulin, and leptin levels of male and female rats exposed to anoxia and control treatments, both at P35 and P95. Furthermore, although at P35, anoxia increased insulin levels of males, it decreased the area of the ß-positive cells in the pancreas of females. At P95, anoxia increased post-prandial weight loss of males, post-fasting food intake, insulin, and leptin, and decreased Fos expression in the arcuate nucleus (ARC) of males and females. Hyperphagia was associated with possible resistance to leptin and insulin, suspected by the high circulating levels of these hormones and poor neuronal activation of ARC. This study demonstrated that continuous social isolation from weaning modifies, in a differentiated way, the long-term energy metabolism and growth of male and female Wistar rats exposed to neonatal anoxia or even control treatments. Therefore, social isolation should be considered as a factor that negatively influences experimental results and the outcomes of the neonatal injury. These results should also be taken into account in clinical procedures, since the used model simulates the preterm babies' conditions and some therapeutic approaches require isolation.

Prenatal exposition to haloperidol: A preclinical narrative review.

Pre-existing maternal mental disorders may affect the early interactions between mother and baby, impacting the child's psychoemotional development. The typical antipsychotic haloperidol can be used during pregnancy, even with some restrictions. Its prescription is not limited to psychotic disorders, but also to other psychiatric conditions of high incidence and prevalence in the woman's fertile period. The present review focused on the preclinical available data regarding the biological and behavioral implications of embryonic exposure to haloperidol. The understanding of the effects of psychotropic drugs during neurodevelopment is important for its clinical aspect since there is limited evidence regarding the risks of antipsychotic drug treatment in pregnant women and their children. Moreover, a better comprehension of the mechanistic events that can be affected by antipsychotic treatment during the critical period of neurodevelopment may offer insights into the pathophysiology of neurodevelopmental disorders. The findings presented in this review converge to the existence of several risks associated with prenatal exposure to such medication and emphasize the need for further studies regarding its dimensions.

The Impact of Hydroxytyrosol on the Metallomic-Profile in an Animal Model of Alzheimer's Disease.

It is undeniable that as people get older, they become progressively more susceptible to neurodegenerative illnesses such as Alzheimer's disease (AD). Memory loss is a prominent symptom of this condition and can be exacerbated by uneven levels of certain metals. This study used inductively coupled plasma mass spectrometry (ICP-MS) to examine the levels of metals in the blood plasma, frontal cortex, and hippocampus of Wistar rats with AD induced by streptozotocin (STZ). It also tested the effects of the antioxidant hydroxytyrosol (HT) on metal levels. The Barnes maze behavior test was used, and the STZ group showed less certainty and greater distance when exploring the Barnes maze than the control group. The results also indicated that the control group and the STZ + HT group exhibited enhanced learning curves during the Barnes maze training as compared to the STZ group. The ICP-MS analysis showed that the STZ group had lower levels of cobalt in their blood plasma than the control group, while the calcium levels in the frontal cortex of the STZ + HT group were higher than in the control group. The most important finding was that copper levels in the frontal cortex from STZ-treated animals were higher than in the control group, and that the STZ + HT group returned to equivalent levels to the control group. The antioxidant HT can restore copper levels to their basal physiological state. This finding may help explain HT's potential beneficial effect in AD-patients.

Consequences of oxygen deprivation on myelination and sex-dependent alterations.

Oxygen deprivation is one of the main causes of morbidity and mortality in newborns, occurring with a higher prevalence in preterm infants, reaching 20 % to 50 % mortality in newborns in the perinatal period. When they survive, 25 % exhibit neuropsychological pathologies, such as learning difficulties, epilepsy, and cerebral palsy. White matter injury is one of the main features found in oxygen deprivation injury, which can lead to long-term functional impairments, including cognitive delay and motor deficits. The myelin sheath accounts for much of the white matter in the brain by surrounding axons and enabling the efficient conduction of action potentials. Mature oligodendrocytes, which synthesize and maintain myelination, also comprise a significant proportion of the brain's white matter. In recent years, oligodendrocytes and the myelination process have become potential therapeutic targets to minimize the effects of oxygen deprivation on the central nervous system. Moreover, evidence indicate that neuroinflammation and apoptotic pathways activated during oxygen deprivation may be influenced by sexual dimorphism. To summarize the most recent research about the impact of sexual dimorphism on the neuroinflammatory state and white matter injury after oxygen deprivation, this review presents an overview of the oligodendrocyte lineage development and myelination, the impact of oxygen deprivation and neuroinflammation on oligodendrocytes in neurodevelopmental disorders, and recent reports about sexual dimorphism regarding the neuroinflammation and white matter injury after neonatal oxygen deprivation.

The impact of antidepressants on human neurodevelopment: Brain organoids as experimental tools.

The use of antidepressants during pregnancy benefits the mother's well-being, but the effects of such substances on neurodevelopment remain poorly understood. Moreover, the consequences of early exposure to antidepressants may not be immediately apparent at birth. In utero exposure to selective serotonin reuptake inhibitors (SSRIs) has been related to developmental abnormalities, including a reduced white matter volume. Several reports have observed an increased incidence of autism spectrum disorder (ASD) and attention deficit hyperactivity disorder (ADHD) after prenatal exposure to SSRIs such as sertraline, the most widely prescribed SSRI. The advent of human-induced pluripotent stem cell (hiPSC) methods and assays now offers appropriate tools to test the consequences of such compounds for neurodevelopment in vitro. In particular, hiPSCs can be used to generate cerebral organoids - self-organized structures that recapitulate the morphology and complex physiology of the developing human brain, overcoming the limitations found in 2D cell culture and experimental animal models for testing drug efficacy and side effects. For example, single-cell RNA sequencing (scRNA-seq) and electrophysiological measurements on organoids can be used to evaluate the impact of antidepressants on the transcriptome and neuronal activity signatures in developing neurons. While the analysis of large-scale transcriptomic data depends on dimensionality reduction methods, electrophysiological recordings rely on temporal data series to discriminate statistical characteristics of neuronal activity, allowing for the rigorous analysis of the effects of antidepressants and other molecules that affect the developing nervous system, especially when applied in combination with relevant human cellular models such as brain organoids.

Impact of neonatal anoxia and hypothermic treatment on development and memory of rats.

Therapeutic hypothermia (TH) is well established as a standard treatment for term and near-term infants. However, therapeutic effects of hypothermia following neonatal anoxia in very premature babies remains inconclusive. The present rodent model of preterm neonatal anoxia has been shown to alter developmental milestones and hippocampal neurogenesis, and to disrupt spatial learning and memory in adulthood. These effects seem to be reduced by post-insult hypothermia. Epigenetic-related mechanisms have been postulated as valuable tools for developing new therapies. Dentate gyrus neurogenesis is regulated by epigenetic factors. This study evaluated whether TH effects in a rodent model of preterm oxygen deprivation are based on epigenetic alterations. The effects of TH on both developmental features (somatic growth, maturation of physical characteristics and early neurological reflexes) and performance of behavioral tasks at adulthood (spatial reference and working memory, and fear conditioning) were investigated in association with the possible involvement of the epigenetic operator Enhancer of zeste homolog 2 (Ezh2), possibly related to long-lasting effects on hippocampal neurogenesis. Results showed that TH reduced both anoxia-induced hippocampal neurodegeneration and anoxia-induced impairments on risk assessment behavior, acquisition of spatial memory, and extinction of auditory and contextual fear conditioning. In contrast, TH did not prevent developmental alterations caused by neonatal anoxia and did not restore hippocampal neurogenesis or cause changes in EZH2 levels. In conclusion, despite the beneficial effects of TH in hippocampal neurodegeneration and in reversing disruption of performance of behavioral tasks following oxygen deprivation in prematurity, these effects seem not related to developmental alterations and hippocampal neurogenesis and, apparently, is not caused by Ezh2-mediated epigenetic alteration.

Small Molecule GSK-J1 Affects Differentiation of Specific Neuronal Subtypes in Developing Rat Retina.

Histone post-translational modification has been shown to play a pivotal role in regulating gene expression and fate determination during the development of the central nervous system. Application of pharmacological blockers that control histone methylation status has been considered a promising avenue to control abnormal developmental processes and diseases as well. In this study, we focused on the role of potent histone demethylase inhibitor GSK-J1 as a blocker of Jumonji domain-containing protein 3 (Jmjd3) in early postnatal retinal development. Jmjd3 participates in different processes such as cell proliferation, apoptosis, differentiation, senescence, and cell reprogramming via demethylation of histone 3 lysine 27 trimethylation status (H3K27 me3). As a first approach, we determined the localization of Jmjd3 in neonate and adult rat retina. We observed that Jmjd3 accumulation is higher in the adult retina, which is consistent with the localization in the differentiated neurons, including ganglion cells in the retina of neonate rats. At this developmental age, we also observed the presence of Jmjd3 in undifferentiated cells. Also, we confirmed that GSK-J1 caused the increase in the H3k27 me3 levels in the retinas of neonate rats. We next examined the functional consequences of GSK-J1 treatment on retinal development. Interestingly, injection of GSK-J1 simultaneously increased the number of proliferative and apoptotic cells. Furthermore, an increased number of immature cells were detected in the outer plexiform layer, with longer neuronal processes. Finally, the influence of GSK-J1 on postnatal retinal cytogenesis was examined. Interestingly, GSK-J1 specifically caused a significant decrease in the number of PKCα-positive cells, which is a reliable marker of rod-on bipolar cells, showing no significant effects on the differentiation of other retinal subtypes. To our knowledge, these data provide the first evidence that in vivo pharmacological blocking of histone demethylase by GSK-J1 affects differentiation of specific neuronal subtypes. In summary, our results indisputably revealed that the application of GSK-J1 could influence cell proliferation, maturation, apoptosis induction, and specific cell determination. With this, we were able to provide evidence that this small molecule can be explored in therapeutic strategies for the abnormal development and diseases of the central nervous system.

Behavioral, cognitive and histological changes following neonatal anoxia: Male and female rats' differences at adolescent age.

Neonatal anoxia induces long-term brain injury that may underlie neurobehavioral deficits at adolescence. Neonatal anoxia, induced by exposure of 30-hour old pups to 100% nitrogen, represents a non-invasive and global stimulus, which simulates clinical conditions of human pre-term babies (around 6 gestational months). Previous studies showed that neonatal anoxia induced impairments of spatial memory and altered anxiety-like behaviors in male rats tested at adult age. This study evaluated if neonatal anoxia induces similar behavioral effects in female rats, as compared to males, by testing the animals at adolescence, and also searched for possible cell losses in hippocampal subfields. Results in the Elevated Plus Maze test showed that anoxic females spent proportionally more time within the open arms as compared to anoxic males, suggesting a less anxious-like behavior. In the Morris Water Maze Test, latencies and path lengths of the anoxic subjects were longer as compared to control subjects, thus indicating that anoxia disrupted the cognitive functions required for spatial mapping. In addition, results showed that anoxia-induced disruption was greater in male rats as compared to female rats. Stereological analysis revealed that anoxic male rats exhibited significant cell losses in the dorsal hippocampus dentate gyrus and CA1 subfields, but not in CA3-2 subfield. Similar results were observed in the ventral hippocampus, but now with cell loss in the male CA3-2 subfield. There were also significant cell loss differences of anoxic male rats as compared to anoxic female rats. In conclusion, neonatal anoxia induces deleterious and long lasting behavioral and cognitive disruptions, and these effects were stronger in male rats as compared to female rats. These changes are congruent with the pattern of cell losses observed in hippocampal subfields. Together, these results emphasize the relevance of scientific research, aiming at clinical strategies and treatments, consider the sex differential patterns of response to neonatal injury.

Microbiota influence the development of the brain and behaviors in C57BL/6J mice.

We investigated the contributions of commensal bacteria to brain structural maturation by magnetic resonance imaging and behavioral tests in four and 12 weeks old C57BL/6J specific pathogen free (SPF) and germ free (GF) mice. SPF mice had increased volumes and fractional anisotropy in major gray and white matter areas and higher levels of myelination in total brain, major white and grey matter structures at either four or 12 weeks of age, demonstrating better brain maturation and organization. In open field test, SPF mice had better mobility and were less anxious than GF at four weeks. In Morris water maze, SPF mice demonstrated better spatial and learning memory than GF mice at 12 weeks. In fear conditioning, SPF mice had better contextual memory than GF mice at 12 weeks. In three chamber social test, SPF mice demonstrated better social novelty than GF mice at 12 weeks. Our data demonstrate numerous significant differences in morphological brain organization and behaviors between SPF and GF mice. This suggests that commensal bacteria are necessary for normal morphological development and maturation in the grey and white matter of the brain regions with implications for behavioral outcomes such as locomotion and cognitive functions.

Evaluation of Possible Consequences of Zika Virus Infection in the Developing Nervous System.

The Zika virus (ZIKV) outbreak that occurred in the northeast of Brazil in 2015 led to alarming numbers of babies born with microcephaly in this region. Since then, several studies have evaluated the relationship between ZIKV infection and development of the malformation although the specific mechanistic interaction between ZIKV and human physiological processes that ultimately manifest as microcephaly remains debated. Importantly, most current studies did not consider the specificities of the biology and life cycle of ZIKV. As a consequence, specificities of the infection on the developing central nervous system (CNS) were frequently disregarded. In order to begin to address this important gap in our knowledge, we have collated and critically reviewed the existing evidence in this area to identify any emerging consensus on this topic and thereafter describe possible mechanisms by which ZIKV infection could interfere with specific processes of CNS development, such as neuronal proliferation, and the complex interactions of immature neurons with radial glial cells. With this, we were able to present the current knowledge on this important topic in the neurobiology field.

Sex differences in somatic and sensory motor development after neonatal anoxia in Wistar rats.

Currently, one of the important causes of brain injury in new-borns is the neonatal anoxia which impacts the perinatology services worldwide. Animal models of anoxia have been used to assess its effects at cellular and behavioural levels in all ages, but few studies focus on sex differences. This study aimed to investigate some physical parameters of development, sensorimotor alterations, early neurological reflexes as well as the density of cells in motor and sensorimotor cerebral cortex of adolescent rats submitted to neonatal anoxia. The results presented significant differences in most of the evaluated parameters, such as body weight and lenght, medio-lateral head axis, eruption of superior incisor, palmar grasp, auditory startle, negative geotaxis, showing that neonatal anoxia affects physical parameters and neurological development, with sex differences. Cellular analysis revealed decreased amount of neurons in motor cortex and primary sensory hind limb and forelimb regions in anoxic group, along with gender difference, as compared to control groups. There is an important rationale for performing early assessment of sensorimotor deficits as there is similarity of the model with high risk human neonates and the sequelae in later life periods, which can be inferred from the present results with suggestion of a possible correlation between sensorimotor development delay and cellular changes in sensorimotor cortex. Furthermore, these observed sex dependent alterations certainly will address further studies and should be considered especially in treatments and strategies to avoid or minimize the neonatal anoxic effects.

Functional Role of Intracellular Calcium Receptor Inositol 1,4,5-Trisphosphate Type 1 in Rat Hippocampus after Neonatal Anoxia.

Anoxia is one of the most prevalent causes of neonatal morbidity and mortality, especially in preterm neonates, constituting an important public health problem due to permanent neurological sequelae observed in patients. Oxygen deprivation triggers a series of simultaneous cascades, culminating in cell death mainly located in more vulnerable metabolic brain regions, such as the hippocampus. In the process of cell death by oxygen deprivation, cytosolic calcium plays crucial roles. Intracellular inositol 1,4,5-trisphosphate receptors (IP3Rs) are important regulators of cytosolic calcium levels, although the role of these receptors in neonatal anoxia is completely unknown. This study focused on the functional role of inositol 1,4,5-trisphosphate receptor type 1 (IP3R1) in rat hippocampus after neonatal anoxia. Quantitative real-time PCR revealed a decrease of IP3R1 gene expression 24 hours after neonatal anoxia. We detected that IP3R1 accumulates specially in CA1, and this spatial pattern did not change after neonatal anoxia. Interestingly, we observed that anoxia triggers translocation of IP3R1 to nucleus in hippocampal cells. We were able to observe that anoxia changes distribution of IP3R1 immunofluorescence signals, as revealed by cluster size analysis. We next examined the role of IP3R1 in the neuronal cell loss triggered by neonatal anoxia. Intrahippocampal injection of non-specific IP3R1 blocker 2-APB clearly reduced the number of Fluoro-Jade C and Tunel positive cells, revealing that activation of IP3R1 increases cell death after neonatal anoxia. Finally, we aimed to disclose mechanistics of IP3R1 in cell death. We were able to determine that blockade of IP3R1 did not reduced the distribution and pixel density of activated caspase 3-positive cells, indicating that the participation of IP3R1 in neuronal cell loss is not related to classical caspase-mediated apoptosis. In summary, this study may contribute to new perspectives in the investigation of neurodegenerative mechanisms triggered by oxygen deprivation.

Determining the Roles of Inositol Trisphosphate Receptors in Neurodegeneration: Interdisciplinary Perspectives on a Complex Topic.

It is well known that calcium (Ca2+) is involved in the triggering of neuronal death. Ca2+ cytosolic levels are regulated by Ca2+ release from internal stores located in organelles, such as the endoplasmic reticulum. Indeed, Ca2+ transit from distinct cell compartments follows complex dynamics that are mediated by specific receptors, notably inositol trisphosphate receptors (IP3Rs). Ca2+ release by IP3Rs plays essential roles in several neurological disorders; however, details of these processes are poorly understood. Moreover, recent studies have shown that subcellular location, molecular identity, and density of IP3Rs profoundly affect Ca2+ transit in neurons. Therefore, regulation of IP3R gene products in specific cellular vicinities seems to be crucial in a wide range of cellular processes from neuroprotection to neurodegeneration. In this regard, microRNAs seem to govern not only IP3Rs translation levels but also subcellular accumulation. Combining new data from molecular cell biology with mathematical modelling, we were able to summarize the state of the art on this topic. In addition to presenting how Ca2+ dynamics mediated by IP3R activation follow a stochastic regimen, we integrated a theoretical approach in an easy-to-apply, cell biology-coherent fashion. Following the presented premises and in contrast to previously tested hypotheses, Ca2+ released by IP3Rs may play different roles in specific neurological diseases, including Alzheimer's disease and Parkinson's disease.

Impact of neonatal anoxia on adult rat hippocampal volume, neurogenesis and behavior.

Neonates that suffer oxygen deprivation during birth can have long lasting cognitive deficits, such as memory and learning impairments. Hippocampus, one of the main structures that participate in memory and learning processes, is a plastic and dynamic structure that conserves during life span the property of generating new cells which can become neurons, the so-called neurogenesis. The present study investigated whether a model of rat neonatal anoxia, that causes only respiratory distress, is able to alter the hippocampal volume, the neurogenesis rate and has functional implications in adult life. MRI analysis revealed significant hippocampal volume decrease in adult rats who had experienced neonatal anoxia compared to control animals for rostral, caudal and total hippocampus. In addition, these animals also had 55.7% decrease of double-labelled cells to BrdU and NeuN, reflecting a decrease in neurogenesis rate. Finally, behavioral analysis indicated that neonatal anoxia resulted in disruption of spatial working memory, similar to human condition, accompanied by an anxiogenic effect. The observed behavioral alterations caused by oxygen deprivation at birth might represent an outcome of the decreased hippocampal neurogenesis and volume, evidenced by immunohistochemistry and MRI analysis. Therefore, based on current findings we propose this model as suitable to explore new therapeutic approaches.

Interplay Between Exosomes, microRNAs and Toll-Like Receptors in Brain Disorders.

Extracellular vesicles (EVs), including exosomes, microvesicles and apoptotic bodies, participate in intercellular communication, and particularly, in paracrine and endocrine signalling. The EVs and their specific contents have been considered hallmarks of different diseases. It has been recently discovered that EVs can co-transport nucleic acids such as DNAs, ribosomal RNAs, circular RNAs (circRNAs), long noncoding RNAs (lnRNAs) and microRNAs (miRNAs). miRNAs are important regulators of gene expression at the post-transcriptional level, although they may also play other roles. Recent evidence supports the hypothesis that miRNAs can activate Toll-like receptors (TLRs) under certain circumstances. TLRs belong to a multigene family of immune system receptors and have been recently described in the nervous system. In the immune system, TLRs are important for the recognition of the invading microorganisms, whereas in the nervous system, they recognise endogenous ligands released by undifferentiated or necrotic/injured cells. In the neuronal disease field, TLRs activity has been associated with amyotrophic lateral sclerosis (ALS), stroke, Alzheimer's and Parkinson's disease. Herein, we reviewed the current knowledge of the relationship between miRNA release by EVs and the inflammation signalling triggered by TLRs in neighbouring cells or during long-distance cell-to-cell communication. We highlight novel aspects of this communication mechanism, offering a valuable insight into such pathways in health and disease.

Spatial learning and neurogenesis: Effects of cessation of wheel running and survival of novel neurons by engagement in cognitive tasks.

Physical exercise stimulates cell proliferation in the adult dentate gyrus and facilitates acquisition and/or retention of hippocampal-dependent tasks. It is established that regular physical exercise improves cognitive performance. However, it is unclear for how long these benefits last after its interruption. Independent groups of rats received both free access to either unlocked (EXE Treatment) or locked (No-EXE Treatment) running wheels for 7 days, and daily injections of bromodeoxyuridine (BrdU) in the last 3 days. After a time delay period of either 1, 3, or 6 weeks without training, the animals were tested in the Morris water maze (MWM) either in a working memory task dependent on hippocampal function (MWM-HD) or in a visible platform searching task, independent on hippocampal function (MWM-NH). Data confirmed that exposure of rats to 7 days of spontaneous wheel running increases cell proliferation and neurogenesis. In contrast, neurogenesis was not accompanied by significant improvements of performance in the working memory version of the MWM. Longer time delays between the end of exercise and the beginning of cognitive training in the MWM resulted in lower cell survival; that is, the number of novel surviving mature neurons was decreased when this delay was 6 weeks as compared with when it was 1 week. In addition, data showed that while exposure to the MWM-HD working memory task substantially increased survival of novel neurons, exposure to the MWM-NH task did not, thus indicating that survival of novel dentate gyrus neurons depends on the engagement of this brain region in performance of cognitive tasks. © 2015 Wiley Periodicals, Inc.

Elevated spinal monoamine neurotransmitters after antenatal hypoxia-ischemia in rabbit cerebral palsy model.

We hypothesized that a deficiency in the descending serotonergic input to spinal cord may underlie postnatal muscle hypertonia after global antenatal hypoxic-ischemic injury in a rabbit model of cerebral palsy. Neurotransmitter content was determined by HPLC in the spinal cord of newborns with and without muscle hypertonia after fetal global hypoxic-ischemic brain injury and naïve controls. Contrary to our hypothesis, serotonin levels in both cervical and lumbar expansions and norepinephrine in cervical expansion were increased in hypertonic kits relative to non-hypertonic kits and controls, with unchanged number of serotonergic cells in caudal raphe by stereological count. Serotonergic fiber length per unit of volume was also increased in hypertonic kits' cervical and lumbar spinal cord, both in dorsal and ventral horns. Gene expression of serotonin transporter was increased and 5-HTR2 receptors were decreased in hypertonic kits relative to controls in cervical and lumbar cord. Intrathecal administration of non-selective serotonin receptor inhibitor methysergide decreased muscle tone in hypertonic kits only. Conversely, intrathecal administration of serotonin solution increased muscle tone only in non-hypertonic kits. We speculate that maturation of serotonergic system in spinal cord may be directly affected by decreased corticospinal connectivity after antenatal hypoxic-ischemic brain injury. Following prenatal hypoxia-ischemia, newborn rabbits exhibit elevated levels of serotonin in the spinal cord that were linked to muscle hypertonia. Serotonergic terminal density was also increased in hypertonic newborns' spinal cord. Intrathecal administration of the non-selective serotonin receptor inhibitor methysergide decreased muscle tone in hypertonic newborns only. Elevated spinal serotonin thus suggests a novel pathophysiological mechanism of hypertonia in cerebral palsy.

Immunohistochemistry and ultrastructural characteristics of nerve endings in the oral mucosa of rat.

The sensory nerve endings of the rat tongue, cheek and palate were studied using immunohistochemical staining and transmission electron microscopy analysis. The specimens were fixed in modified Karnovsky solution and embedded in Spurr resin. Substance P, calcitonin gene-related peptide (CGRP)- and protein gene product 9.5 (PGP b9.5)-containing nerve fibers in the rat tongue, cheek and palate were examined by electronic microscopical analysis and immunohistochemical localization. These fibers run very close to the basal lamina of the epithelium and extend into the filliform and fungiform papillae. Numerous plexiform fibers immunoreactive for substance P, CGRP and PGP 9.5 were found in the connective tissue of mucosa. Electron microscopic observations showed clearly immunostained nerve fibers, which are located very close to the basal lamina of epithelial cells. Some electron-dense granules may be observed in the axoplasms of both substance P and CGRP immunoreactive fibers. Several lamellar corpuscles into the subepithelial connective tissue papillae, Merkel corpuscles and numerous thin unmyelinated and myelinated axons were observed. The terminal axons revealed numerous mitochondria, neurofilaments, microtubules and clear vesicles in the base of axoplasmic protrusions. The lamellar cells showed caveolae and interlamelar spaces filled by amorphous substance. Between the lamellar cells and axoplasmic membrane, and in the adjacent lamellae region, desmosome-type junctions were observed. The quantitative and morphometric analysis showed nerve endings with an average area of 4.83 ± 3.4 µm(2) and 19.4 internal mitochondria in this site and the organized corpuscles with an average area of 79.24 ± 27.24 µm(2) and 24.23 internal mitochondria in this place. All the structures observed are involved in the transmission of pain and mechanoreceptors stimulus of these oral mucosae.

Near-term fetal hypoxia-ischemia in rabbits: MRI can predict muscle tone abnormalities and deep brain injury.

BACKGROUND AND PURPOSE: The pattern of antenatal brain injury varies with gestational age at the time of insult. Deep brain nuclei are often injured at older gestational ages. Having previously shown postnatal hypertonia after preterm fetal rabbit hypoxia-ischemia, the objective of this study was to investigate the causal relationship between the dynamic regional pattern of brain injury on MRI and the evolution of muscle tone in the near-term rabbit fetus. METHODS: Serial MRI was performed on New Zealand white rabbit fetuses to determine equipotency of fetal hypoxia-ischemia during uterine ischemia comparing 29 days gestation (E29, 92% gestation) with E22 and E25. E29 postnatal kits at 4, 24, and 72 hours after hypoxia-ischemia underwent T2- and diffusion-weighted imaging. Quantitative assessments of tone were made serially using a torque apparatus in addition to clinical assessments. RESULTS: Based on the brain apparent diffusion coefficient, 32 minutes of uterine ischemia was selected for E29 fetuses. At E30, 58% of the survivors manifested hind limb hypotonia. By E32, 71% of the hypotonic kits developed dystonic hypertonia. Marked and persistent apparent diffusion coefficient reduction in the basal ganglia, thalamus, and brain stem was predictive of these motor deficits. CONCLUSIONS: MRI observation of deep brain injury 6 to 24 hours after near-term hypoxia-ischemia predicts dystonic hypertonia postnatally. Torque-displacement measurements indicate that motor deficits in rabbits progressed from initial hypotonia to hypertonia, similar to human cerebral palsy, but in a compressed timeframe. The presence of deep brain injury and quantitative shift from hypo- to hypertonia may identify patients at risk for developing cerebral palsy.