Postnatal Rat Research Articles

Transfer of rhodamine-123\xa0into the brain and cerebrospinal fluid of fetal, neonatal and adult rats

BackgroundAdenosine triphosphate binding cassette transporters such as P-glycoprotein (PGP) play an important role in drug pharmacokinetics by actively effluxing their substrates at barrier interfaces, including the blood-brain, blood-cerebrospinal fluid (CSF) and placental barriers. For a molecule to access the brain during fetal stages it must bypass efflux transporters at both the placental barrier and brain barriers themselves. Following birth, placental protection is no longer present and brain barriers remain the major line of defense. Understanding developmental differences that exist in the transfer of PGP substrates into the brain is important for ensuring that medication regimes are safe and appropriate for all patients.MethodsIn the present study PGP substrate rhodamine-123 (R123) was injected intraperitoneally into E19 dams, postnatal (P4, P14) and adult rats. Naturally fluorescent properties of R123 were utilized to measure its concentration in blood-plasma, CSF and brain by spectrofluorimetry (Clariostar). Statistical differences in R123 transfer (concentration ratios between tissue and plasma ratios) were determined using Kruskal-Wallis tests with Dunn’s corrections.ResultsFollowing maternal injection the transfer of R123 across the E19 placenta from maternal blood to fetal blood was around 20 %. Of the R123 that reached fetal circulation 43 % transferred into brain and 38 % into CSF. The transfer of R123 from blood to brain and CSF was lower in postnatal pups and decreased with age (brain: 43 % at P4, 22 % at P14 and 9 % in adults; CSF: 8 % at P4, 8 % at P14 and 1 % in adults). Transfer from maternal blood across placental and brain barriers into fetal brain was approximately 9 %, similar to the transfer across adult blood-brain barriers (also 9 %). Following birth when placental protection was no longer present, transfer of R123 from blood into the newborn brain was significantly higher than into adult brain (3 fold, p < 0.05).ConclusionsAdministration of a PGP substrate to infant rats resulted in a higher transfer into the brain than equivalent doses at later stages of life or equivalent maternal doses during gestation. Toxicological testing of PGP substrate drugs should consider the possibility of these patient specific differences in safety analysis.

Rutin improves glutamate uptake and inhibits glutamate excitotoxicity in rat brain slices.

Rutin is an important flavonoid consumed in the daily diet. It is also known as vitamin P and has been extensively investigated due to its pharmacological properties. On the other hand, neuronal death induced by glutamate excitotoxicity is present in several diseases including neurodegenerative diseases. The neuroprotective properties of rutin have been under investigation, although its mechanism of action is still poorly understood. We hypothesized that the mechanisms of neuroprotection of rutin are associated with the increase in glutamate metabolism in astrocytes. This study aimed to evaluate the protective effects of rutin with a focus on the modulation of glutamate detoxification. We used brain organotypic cultures from post-natal Wistar rats (P7-P9) treated with rutin to evaluate neural cell protection and levels of proteins involved in the glutamate metabolism. Moreover, we used cerebral cortex slices from adult Wistar rats to evaluate glutamate uptake. We showed that rutin inhibited the cell death and loss of glutamine synthetase (GS) induced by glutamate that was associated with an increase in glutamate-aspartate transporter (GLAST) in brain organotypic cultures from post-natal Wistar rats. Additionally, it was observed that rutin increased the glutamate uptake in cerebral cortex slices from adult Wistar rats. We conclude that rutin is a neuroprotective agent that prevents glutamate excitotoxicity and thereof suggest that this effect involves the regulation of astrocytic metabolism.

Purinergic Receptor Blockade with Suramin Increases Survival of Postnatal Neural Progenitor Cells In Vitro.

Neural progenitor cells (NPCs) are self-renewing and multipotent cells that persist in the postnatal and adult brain in the subventricular zone and the hippocampus. NPCs can be expanded in vitro to be used in cell therapy. However, expansion is limited, since the survival and proliferation of adult NPCs decrease with serial passages. Many signaling pathways control NPC survival and renewal. Among these, purinergic receptor activation exerts differential effects on the biology of adult NPCs depending on the cellular context. In this study, we sought to analyze the effect of a general blockade of purinergic receptors with suramin on the proliferation and survival of NPCs isolated from the subventricular zone of postnatal rats, which are cultured as neurospheres. Treatment of neurospheres with suramin induced a significant increase in neurosphere diameter and in NPC number attributed to a decrease in apoptosis. Proliferation and multipotency were not affected. Suramin also induced an increase in the gap junction protein connexin43 and in vascular endothelial growth factor, which might be involved in the anti-apoptotic effect. Our results offer a valuable tool for increasing NPC survival before implantation in the lesioned brain and open the possibility of using this drug as adjunctive therapy to NPC transplantation.

Protein expression pattern of calcium-responsive transactivator in early postnatal and adult testes

Calcium-responsive transactivator (CREST), a nuclear protein highly expressed in postmitotic neurons, is involved in the regulation of cell cycle, differentiation and dendritic development of neuronal cells. Its mRNA has been detected in the testis of adult rat, whilst its protein expression and distribution pattern in the testis remain to be elucidated. In this study, we examined the distribution of CREST in the adult testes of both rats and human as well as the expression pattern of CREST in the testes of postnatal developing rats. In the adult testes of both human and rats, immunohistochemical analysis revealed that CREST was selectively distributed in the mature Sertoli cells but not in the spermatogenic cells. In the testes of postnatal developmental rats, CREST was expressed not only in Sertoli cells but also in the gonocytes and spermatogenic cells at the initial stage of spermatogenic cell differentiation. CREST immunoreactivity continued to increase in Sertoli cells during differentiation, reaching its peak in adulthood. However, CREST immunostaining intensity dramatically decreased as the spermatogenic cells differentiate, disappearing in the post-differentiation stage. Furthermore, Brg1 and p300, two CREST-interacting proteins ubiquitously expressed in the body, are found to be colocalized with CREST in the spermatogenic epithelial cells including Sertoli cells. The unique expression pattern of CREST in developing testis suggests that CREST might play regulatory roles in the differentiation of spermatogenic epithelial cells. The Sertoli cell-specific expression of CREST in the adulthood hints that CREST might be a novel biomarker for the mature Sertoli cells.

Blockade of BK channels attenuates chronic visceral hypersensitivity in an IBS-like rat model.

BackgroundVisceral hypersensitivity in irritable bowel syndrome (IBS) is still poorly understood, despite that chronic abdominal pain is the most common symptoms in IBS patients. To study effects of BK channels on visceral hypersensitivity in IBS rats and the underlying mechanisms, IBS rats were established by colorectal distention (CRD) in postnatal rats. The expression of large-conductance calcium and voltage-dependent potassium ion channels (BK channels) of the thoracolumbar spinal cord was examined in IBS and control rats. The effects of BK channel blockade on visceral hypersensitivity were evaluated. The interaction of BK channels and N-methyl-D-aspartate acid (NMDA) receptors was explored, and synaptic transmission at superficial dorsal horn (SDH) neurons of the thoracolumbar spinal cord was recorded by whole-cell patch clamp in IBS rats.ResultsThe expression of the BK channels of the thoracolumbar spinal cord in IBS rats was significantly reduced. The blockade of BK channels could reduce the visceral hypersensitivity in IBS rats. There was an interaction between BK channels and NMDA receptors in the spinal cord. The frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) in SDH neurons is significantly reduced in IBS rats. The blockade of BK channels depolarizes the inhibitory interneuron membrane and increases their excitability in IBS rats.ConclusionsBK channels could interact with NMDA receptors in the thoracolumbar spinal cord of rats and regulate visceral hypersensitivity in IBS rats.

Developmental Expression of Cyclooxygenase isoforms in the hepatocytes of Albino Rat

AbstractBackground: The liver represents the major organ that participates in the elimination and degradation of arachidonic acid of systemic origin. This study is one of few types of research that studied the detailed expression of cyclooxygenase in the hepatocytes and other cells in liver tissue at different stages of maturation. Material and methods: A total number of 8 embryos and 40 rats at different ages postnatal were used. The prenatal rats were 18 days old of gestation while the postnatal rats were at 1, 7, 14, 21, and 28 days old. Aim of the work: The present study is concerned with exploring and compare the developmental expression of cyclooxygenase (COX-1, COX-2, and COX-3) in the liver tissue using immunohistochemical techniques. This will open the future for a new rationale of the optimal therapeutic use of COX in liver diseases.Results: The results showed that, in liver cells, the expression of COX isoforms was scientifically changed with the development of the liver tissue. COX-1 and COX-2 immunostaining were noticed to be the same in some ages however, the COX-1 expression resembled the COX-3 expression in other ages. Conclusion: It can conclude the expression of three types of COX isoforms in liver tissue played an important role in the development and maturation of different cells population either through prostaglandin production or a novel gene production. Nuclear localization of COX in the hepatocytes arises a lot of questions about the role of COX in the enhancement of liver division and maturation.

Development of myodural bridge located within the atlanto-occipital interspace of rats.

The myodural bridge (MDB) is a dense connective tissue structure that connects the subocciptal musculature to the spinal dura mater. The purpose of this study was to clarify morphological evolution characteristics and compositional changes in the fibrous structures of MDB during its growth and development in the atlanto-occipital interspace. For this, histological sections from Sprague-Dawley (SD) rats (age, E17 to adulthood) were stained with Masson's Trichrome and Picrosirius Red. The results demonstrated that at age E18, the posterior arch of the atlas was completely closed and MDB fibers had already begun to form. In rat embryos (E18-E21), only few fibers and muscles were present in the suboccipital region, and these were lightly stained. In postnatal rats, an obvious increase in the amount of fibers and muscle tissues was noted. At age P1, MDB fibers originated from the rectus capitis posterior minor muscle and merged into the atlanto-occipital membrane, which was closely attached to the spinal dura mater. As rats matured, MDB fibers gradually became denser and more organized. This study also showed that in postnatal rats, MDB was mainly composed of type I collagen fibers. By observing the development of MDB in SD rats, the function of MDB can be further understood. This study provides a morphological basis for future functional studies involving the MDB.

Hyperexcitability of the Nucleus Accumbens Is Involved in Noise-Induced Hyperacusis.

Reduced tolerance to sound stimuli (hyperacusis) is commonly seen in tinnitus patients. Dysfunction of limbic systems, such as the nucleus accumbens (NAc), may be involved in emotional reactions to the sound stimuli in tinnitus patients. To study the functional changes in the NAc in hyperacusis, we have examined the neural activity changes of the NAc using c-Fos staining in an animal model of hyperacusis. The c-Fos staining was also examined in the medial geniculate nucleus (MGN), a central auditory pathway which has neural projections to the NAc. Postnatal rats (14 days) were exposed to loud noise (115 dB SPL, 4 hours for two consecutive days) to induce hyperacusis (n = 4). Rats without noise exposure were used as the controls (n = 4). After P35, rats in both groups were put in a behavioral training for sound detection. After they were trained to detect sound stimuli, their reaction time to noise bursts centered at 2 kHz (40-110 dB SPL) was measured. Rats in the noise group showed a significantly shorter reaction time than those in the control group to the noise bursts at high intensities, suggesting the noise exposure induced hyperacusis behavior. The c-Fos expressions in the NAc and the MGNs of the noise group were significantly higher than those of the control group. Our results suggested that early-age noise exposure caused hyperactivity in the NAc and the MGNs which may induce the loudness increase in these rats.

Validation of prenatal versus postnatal valproic acid rat models of autism: A behavioral and neurobiological study

Despite the increasing prevalence of autism spectrum disorder (ASD), there is still a deficiency in understanding its exact pathophysiology and treatment, therefore validation of translational ASD animal model is warranted. Although strong evidences support the valproic acid (VPA) model of autism, yet a controversy exists regarding the best timing of exposure whether prenatal or postnatal. Accordingly, this study was designed to compare the time dependent effects of VPA exposure as regard its ability to induce autistic like changes in male Wistar rats. In this study, two different protocols of VPA exposure (prenatal and postnatal) were compared at different levels (behavioral, neurochemical and histopathological). Results of this study revealed that both prenatal and postnatal VPA exposures induced autistic-like behaviors manifested by reduced social interaction, increased repetitive stereotyped behavior and anxiety, cognitive dysfunction, lowered sensitivity to pain, and neurodevelopmental delay. Furthermore, inflammatory cytokines and oxidative/nitrosative stress markers were elevated in prefrontal cortex and hippocampal homogenates. Likewise, histopathological and immunohistochemical assessment confirmed the neurodegenerative and the apoptotic changes in prefrontal cortex, hippocampus and cerebellum exhibited by decreased viable cells number and Nissl's granules optical density, and increased caspase-3 immunoreactivity respectively. Interestingly, ASD core symptoms and histopathological changes were significantly (P < 0.05) altered in prenatal VPA model compared to postnatal VPA model. Additionally, postnatal mortality in prenatal model (4.3%) was much lower compared to the postnatal model (22.7%). In conclusion, our study overweighs the ability of prenatal VPA model over postnatal VPA model to induce behavioral and neuropathological alterations that simulate those observed in autistic individuals with a lower postnatal animal mortality, highlighting the privilege of prenatal over postnatal VPA exposure as a translational model for understanding pathophysiology and developing novel targets for management of ASD

Hypothyroidism Induces Interleukin-1-Dependent Autophagy Mechanism as a Key Mediator of Hippocampal Neuronal Apoptosis and Cognitive Decline in Postnatal Rats.

Thyroid hormone (TH) is essential for brain development, and hypothyroidism induces cognitive deficits in children and young adults. However, the participating mechanisms remain less explored. Here, we examined the molecular mechanism, hypothesizing the involvement of a deregulated autophagy and apoptosis pathway in hippocampal neurons that regulate cognitive functions. Therefore, we used a rat model of developmental hypothyroidism, generated through methimazole treatment from gestation until young adulthood. We detected that methimazole stimulated the autophagy mechanism, characterized by increased LC3B-II, Beclin-1, ATG7, and ATG5-12 conjugate and decreased p-mTOR/mTOR and p-ULK1/ULK1 autophagy regulators in the hippocampus of developing and young adult rats. This methimazole-induced hippocampal autophagy could be inhibited by thyroxine treatment. Subsequently, probing the upstream mediators of autophagy revealed an increased hippocampal neuroinflammation, marked by upregulated interleukin (IL)-1alpha and beta and activated microglial marker, Iba1, promoting neuronal IL-1 receptor-1 expression. Hence, IL-1R-antagonist (IL-1Ra), which reduced hippocampal neuronal IL-1R1, also inhibited the enhanced autophagy in hypothyroid rats. We then linked these events with hypothyroidism-induced apoptosis and loss of hippocampal neurons, where we observed that like thyroxine, IL-1Ra and autophagy inhibitor, 3-methyladenine, reduced the cleaved caspase-3 and TUNEL-stained apoptotic neurons and enhanced Nissl-stained neuronal count in methimazole-treated rats. We further related these molecular results with cognition through Y-maze and passive avoidance tests, demonstrating an IL-1Ra and 3-methyladenine-mediated improvement in learning-memory performances of the hypothyroid rats. Taken together, our study enlightens the critical role of neuroinflammation-dependent autophagy mechanism in TH-regulated hippocampal functions, disrupted in developmental hypothyroidism.

Glycyrrhizin Blocks the Detrimental Effects of HMGB1 on Cortical Neurogenesis After Traumatic Neuronal Injury.

Despite medical advances, neurological recovery after severe traumatic brain injury (TBI) remains poor. Elevated levels of high mobility group box protein-1 (HMGB1) are associated with poor outcomes; likely via interaction with receptors for advanced-glycation-end-products (RAGE). We examined the hypothesis that HMGB1 post-TBI is anti-neurogenic and whether this is pharmacologically reversible. Post-natal rat cortical mixed neuro-glial cell cultures were subjected to needle-scratch injury and examined for HMGB1-activation/neuroinflammation. HMGB1-related genes/networks were examined using genome-wide RNA-seq studies in cortical perilesional tissue samples from adult mice. Post-natal rat cortical neural stem/progenitor cell cultures were generated to quantify effects of injury-condition medium (ICM) on neurogenesis with/without RAGE antagonist glycyrrhizin. Needle-injury upregulated TNF-α/NOS-2 mRNA-expressions at 6 h, increased proportions of activated microglia, and caused neuronal loss at 24 h. Transcriptome analysis revealed activation of HMGB1 pathway genes/canonical pathways in vivo at 24 h. A 50% increase in HMGB1 protein expression, and nuclear-to-cytoplasmic translocation of HMGB1 in neurons and microglia at 24 h post-injury was demonstrated in vitro. ICM reduced total numbers/proportions of neuronal cells, but reversed by 0.5 μM glycyrrhizin. HMGB1 is activated following in vivo post mechanical injury, and glycyrrhizin alleviates detrimental effects of ICM on cortical neurogenesis. Our findings highlight glycyrrhizin as a potential therapeutic agent post-TBI.

A novel xeno-organoid approach: exploring the crosstalk between human iPSC-derived PGC-like and rat testicular cells

Specification of germ cell-like cells from induced pluripotent stem cells has become a clinically relevant tool for research. Research on initial embryonic processes is often limited by the access to foetal tissue, and in humans, the molecular events resulting in primordial germ cell (PGC) specification and sex determination remain to be elucidated. A deeper understanding of the underlying processes is crucial to describe pathomechanisms leading to impaired reproductive function. Several protocols have been established for the specification of human pluripotent stem cell towards early PGC-like cells (PGCLC), currently representing the best model to mimic early human germline developmental processes in vitro. Further sex determination towards the male lineage depends on somatic gonadal cells providing the necessary molecular cues. By establishing a culture system characterized by the re-organization of somatic cells from postnatal rat testes into cord-like structures and optimizing efficient PGCLC specification protocols, we facilitated the co-culture of human germ cell-like cells within a surrogate testicular microenvironment. Specified conditions allowed the survival of rat somatic testicular and human PGCLCs for 14 days. Human cells maintained the characteristic expression of octamer-binding transcription factor 4, SRY-box transcription factor 17, and transcription factor AP-2 gamma and were recovered from the xeno-organoids by cell sorting. This novel xeno-organoid approach will allow the in vitro exploration of early sex determination of human PGCLCs.

Dexmedetomidine alleviates neurobehavioral impairments and myelination deficits following lipopolysaccharide exposure in early postnatal rats

AimsWhite matter injury (WMI) is the main form of brain injury in preterm neonate survivors, and perinatal inflammation is implicated in the pathogenesis of WMI. It has been demonstrated that dexmedetomidine, an anesthetic adjuvant, possesses neuroprotective effects in both preclinical and clinical trials. The present study was conducted to explore whether dexmedetomidine could protect against neurobehavioral impairments and myelination deficits caused by lipopolysaccharide (LPS) exposure in the early postnatal rat brain. Main methodsLPS (2 mg/kg) was intraperitoneally (i.p.) injected in Sprague–Dawley rat pups on postnatal day 2 (P2). Dexmedetomidine (25 μg/kg) or vehicle was given i.p. immediately after LPS injection. STAT3 and p-STAT3 expression were detected by western blot in rat brain 24 h after drug administration. Immunostaining for GFAP to was performed to evaluate astrocytic response at 24 h post-LPS and P14. Neurobehavioral tests (the righting reflex, negative geotaxis, and wire hanging maneuver tests) were performed from P5 to P10. Histological analysis of myelin content was accessed by immunohistochemistry for CNPase and MBP at P14. Key findingsOur results showed that treatment with dexmedetomidine significantly ameliorated LPS-induced neurobehavioral abnormalities and myelin damage, which is accompanied by suppression of STAT3 activation and reactive astrogliosis. SignificanceDexmedetomidine can alleviate neurobehavioral impairments and myelination deficits after LPS exposure in early postnatal rats, probably by mitigating STAT3-mediated reactive astrogliosis. Our results suggest that dexmedetomidine might be a promising agent to treat brain injury in neonates.

Effects of early postnatal sevoflurane exposure on oligodendrocyte maturation and myelination in cerebral white matter of the rat

General anaesthesia may impose significant neurocognitive risks on the developing brain. As brain injury in preterm neonates has a particular predilection for cerebral white matter, we aimed to evaluate the effects of sevoflurane on oligodendrocyte maturation and myelination in a preterm-equivalent rat model. Two-day-old postnatal (P2) Sprague-Dawley rats were exposed to 3.3 % (approximately 1 minimum alveolar concentration [MAC]) or 4.9 % (approximately 1.5 MAC) sevoflurane for 2 h. Physiologic parameters were measured at the end of sevoflurane anaesthesia. Oligodendrocyte proliferation, maturation, and myelination were evaluated by immunofluorescence with specific markers at different time points. Open field test and Morris water maze tests were performed to access behavior changes from P29 to P36. Arterial blood gases values and blood glucose levels were within the normal physiologic range. As compared to control, 4.9 %, but not 3.3 % sevoflurane disturbed oligodendrocyte maturation at P14, resulting in hypomyelination and axonal damage in cerebral white matter at P28. Rats exposed to 4.9 %, but not 3.3 % sevoflurane showed decreased explorative activity and increased anxiety-like behaviour, as well as learning and memory impairments. Furthermore, 4.9 %, but not 3.3 % sevoflurane inhibited oligodendrocyte proliferation in the developing white matter of the rat brain at 12 h post-anaesthesia, with further evidence of widespread reactive astrogliosis. High concentration of sevoflurane (4.9 %) exposure in early postnatal rats may disrupt oligodendrocyte maturation and myelination. Our study has aimed a spotlight on the need for safe and rational use of analgesics in neonates, especially preterm infants.

Oleamide Induces Cell Death in Glioblastoma RG2 Cells by a Cannabinoid Receptor-Independent Mechanism.

The endocannabinoid system has been associated with antiproliferative effects in several types of tumors through cannabinoid receptor-mediated cell death mechanisms. Oleamide (ODA) is a CB1/CB2 agonist associated with cell growth and migration by adhesion and/or ionic signals associated with Gap junctions. Antiproliferative mechanisms related to ODA remain unknown. In this work, we evaluated the effects of ODA on cell viability and morphological changes in a rat RG2 glioblastoma cell line and compared these effects with primary astrocyte cultures from 8-day postnatal rats. RG2 and primary astrocyte cultures were treated with ODA at increasing concentrations (25, 50, 100, and 200μM) for different periods of time (12, 24, and 48h). Changes in RG2 cell viability and morphology induced by ODA were assessed by viability/mitochondrial activity test and phase contrast microscopy, respectively. The ratios of necrotic and apoptotic cell death, and cell cycle alterations, were evaluated by flow cytometry. The roles of CB1 and CB2 receptors on ODA-induced changes were explored with specific receptor antagonists. ODA (100μM) induced somatic damage, detachment of somatic bodies, cytoplasmicpolarization, and somatic shrinkage in RG2 cells at 24 and 48h. In contrast, primary astrocytes treated at the same ODA concentrations exhibited cell aggregation but not cell damage. ODA (100μM) increased apoptotic cell death and cell arrest in the G1 phase at 24h in the RG2 line. The effects induced by ODA on cell viability of RG2 cells were independent of CB1 and CB2 receptors or changes in intracellular calcium transient. Results of this novel study suggest that ODA exerts specific antiproliferative effects on RG2 glioblastoma cells through unconventional apoptotic mechanisms not involving canonical signals.

Alcohol Increases Exosome Release from Microglia to Promote Complement C1q-Induced Cellular Death of Proopiomelanocortin Neurons in the Hypothalamus in a Rat Model of Fetal Alcohol Spectrum Disorders.

Microglia, a type of CNS immune cell, have been shown to contribute to ethanol-activated neuronal death of the stress regulatory proopiomelanocortin (POMC) neuron-producing β-endorphin peptides in the hypothalamus in a postnatal rat model of fetal alcohol spectrum disorders. We determined whether the microglial extracellular vesicle exosome is involved in the ethanol-induced neuronal death of the β-endorphin neuron. Extracellular vesicles were prepared from hypothalamic tissues collected from postnatal rats (both males and females) fed daily with 2.5 mg/kg ethanol or control milk formula for 5 d or from hypothalamic microglia cells obtained from postnatal rats, grown in cultures for several days, and then challenged with ethanol or vehicle for 24 h. Nanoparticle tracking analysis and transmission electron microscopy indicated that these vesicles had the size range and shape of exosomes. Ethanol treatments increased the number and the β-endorphin neuronal killing activity of microglial exosomes both in vivo and in vitro Proteomics analyses of exosomes of cultured microglial cells identified a large number of proteins, including various complements, which were elevated following ethanol treatment. Proteomics data involving complements were reconfirmed using quantitative protein assays. Ethanol treatments also increased deposition of the complement protein C1q in β-endorphin neuronal cells in both in vitro and in vivo systems. Recombinant C1q protein increased while C1q blockers reduced ethanol-induced C3a/b, C4, and membrane attack complex/C5b9 formations; ROS production; and ultimately cellular death of β-endorphin neurons. These data suggest that the complement system involving C1q-C3-C4-membrane attack complex and ROS regulates exosome-mediated, ethanol-induced β-endorphin neuronal death.SIGNIFICANCE STATEMENT Neurotoxic action of alcohol during the developmental period is recognized for its involvement in fetal alcohol spectrum disorders, but the lack of clear understanding of the mechanism of alcohol action has delayed the progress in therapeutic intervention of this disease. Proopiomelanocortin neurons known to regulate stress, energy homeostasis, and immune functions are reported to be killed by developmental alcohol exposure because of activation of microglial immune cells in the brain. While microglia are known to use extracellular vesicles to communicate with neurons for maintaining homeostasis, we show here that ethanol exposure during the developmental period hijacks this system to spread apoptotic factors, including complement protein C1q, to induce the membrane attack complex and reactive super-oxygen species for proopiomelanocortin neuronal killing.

Atrial Natriuretic Peptide Improves Neurite Outgrowth from Spiral Ganglion Neurons In Vitro through a cGMP-Dependent Manner.

The spiral ganglion neurons (SGNs) are the primary afferent neurons in the spiral ganglion (SG), while their degeneration or loss would cause sensorineural hearing loss. As a cardiac-derived hormone, atrial natriuretic peptide (ANP) plays a critical role in cardiovascular homeostasis through binding to its functional receptors (NPR-A and NPR-C). ANP and its receptors are widely expressed in the mammalian nervous system where they could be implicated in the regulation of multiple neural functions. Although previous studies have provided direct evidence for the presence of ANP and its functional receptors in the inner ear, their presence within the cochlear SG and their regulatory roles during auditory neurotransmission and development remain largely unknown. Based on our previous findings, we investigated the expression patterns of ANP and its receptors in the cochlear SG and dissociated SGNs and determined the influence of ANP on neurite outgrowth in vitro by using organotypic SG explants and dissociated SGN cultures from postnatal rats. We have demonstrated that ANP and its receptors are expressed in neurons within the cochlear SG of postnatal rat, while ANP may promote neurite outgrowth of SGNs via the NPR-A/cGMP/PKG pathway in a dose-dependent manner. These results indicate that ANP would play a role in normal neuritogenesis of SGN during cochlear development and represents a potential therapeutic candidate to enhance regeneration and regrowth of SGN neurites.

A lab-on-a-chip model of glaucoma.

AimsWe developed a glaucoma‐on‐a‐chip model to evaluate the viability of retinal ganglion cells (RGCs) against high pressure and the potential effect of neuroprotection.MethodsA three‐layered chip consisting of interconnecting microchannels and culture wells was designed and fabricated from poly‐methyl methacrylate sheets. The bottom surface of the wells was modified by air plasma and coated with different membranes to provide a suitable extracellular microenvironment. RGCs were purified from postnatal Wistar rats by magnetic assisted cell sorting up to 70% and characterized by flow cytometry and immunocytochemistry. The cultured RGCs were exposed to normal (15 mmHg) or elevated pressure (33 mmHg) for 6, 12, 24, 36, and 48 hr, with and without adding brain‐derived neurotrophic factor (BDNF) or a novel BDNF mimetic (RNYK).ResultsMultiple inlet ports allow culture media and gas into the wells under elevated hydrostatic pressure. PDL/laminin formed the best supporting membrane. RGC survival rates were 85%, 78%, 70%, 67%, and 61% under normal pressure versus 40%, 22%, 18%, 12%, and 10% under high pressure at 6, 12, 24, 36, and 48 hr, respectively. BDNF and RNYK separately reduced RGC death rates about twofold under both normal and elevated pressures.ConclusionThis model recapitulated the effects of elevated pressure over relatively short time periods and demonstrated the neuroprotective effects of BDNF and RNYK.

Chondroitin sulfate proteoglycan-5 forms perisynaptic matrix assemblies in the adult rat cortex

Composition of the brain extracellular matrix changes in time as maturation proceeds. Chondroitin sulfate proteoglycan 5 (CSPG-5), also known as neuroglycan C, has been previously associated to differentiation since it shapes neurite growth and synapse forming. Here, we show that this proteoglycan persists in the postnatal rat brain, and its expression is higher in cortical regions with plastic properties, including hippocampus and the medial prefrontal cortex at the end of the second postnatal week. Progressively accumulating after birth, CSPG-5 typically concentrates around glutamatergic and GABAergic terminals in twelve-week old rat hippocampus. CSPG-5-containing perisynaptic matrix rings often appear at the peripheral margin of perineuronal nets. Electron microscopy and analysis of synaptosomal fraction showed that CSPG-5 accumulates around, and is associated to synapses, respectively. In vitro analyses suggest that neurons, but less so astrocytes, express CSPG-5 in rat primary neocortical cultures, and CSPG-5 produced by transfected neuroblastoma cells appear at endings and contact points of neurites. In human subjects, CSPG-5 expression shifts in brain areas of the default mode network of suicide victims, which may reflect an impact in the pathogenesis of psychiatric diseases or support diagnostic power.

Effect of M-Cholinoreceptor Blockade on Somatovisceral Intersystemic Interactions in Newborn Rats after Activation of Cholinoreactive Structures

Interaction of slow-wave rhythmic components of cardiac, respiratory and motor activities was analyzed in non-narcotized neonatal (P1) and early postnatal (P16) rat pups under normal conditions and after activation of cholinoreactive structures. A reversible acetylcholinesterase (AChE) inhibitor physostigmine (eserine) and the M-cholinolytic methacin were used in the experiments. It was established that in intact P1 rats intersystemic interactions are mainly realized via oscillations within the near- and multi-minute ranges. Correlative interactions between the rhythms in the decasecond range that were not involved in integrative processes in P1 rats increased significantly at P16. In rat pups of both ages, changes in the intensity and pattern of motor activity (MA) tended to precede the generation of modulatory oscillations of the respiratory and heart rates. AChE inhibition by physostigmine and subsequent activation of cholinoreactive structures evoked a decrease in the level of intersystemic interactions across all ranges of the modulatory rhythms in rats of both ages. The methacin-evoked blockade of peripheral M-cholinoreceptors (M-ChRs) decreased the level of interactions mediated by the near- and multi-minute rhythms at P1 but augmented them at P16. Physostigmine injection at P1 and P16 exerted no significant effect on cardio-somatomotor interactions, with these interactions in P1 rats being maintained by a wider range of the modulatory rhythms. The same situation was also observed during the physostigmine aftereffect period in all interacting systemic pairs. By contrast, after the blockade of peripheral M-ChRs the existing intersystemic interactions were more difficult to be disrupted in P16 rat pups. The activation of cholinoreactive structures, which occurred against the background of M-ChRs blockade, led to disrupt the coordinating interactions in the “MA–respiratory rate” pair. In P1 rats, these functional relationships did not recover, while in P16 rats they were resumed already during the aftereffect period. The power of modulatory rhythms of activity in the systems studied here was not directly related to intersystemic interactions. Changes in intersystemic interactions were due to heterochrony both in the maturation of these functional systems and the involvement of different cholinoreactive structures in the processes of intersystemic regulation.