Endogenous Opioid Signaling in the Mouse Retina Modulates Pupillary Light Reflex.

Opioid peptides and their receptors are expressed in the mammalian retina; however, little is known about how they might affect visual processing. The melanopsin-expressing intrinsically photosensitive retinal ganglion cells (ipRGCs), which mediate important non-image-forming visual processes such as the pupillary light reflex (PLR), express ß-endorphin-preferring, µ-opioid receptors (MORs). The objective of the present study was to elucidate if opioids, endogenous or exogenous, modulate pupillary light reflex (PLR) via MORs expressed by ipRGCs. MOR-selective agonist [D-Ala2, MePhe4, Gly-ol5]-enkephalin (DAMGO) or antagonist D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2 (CTAP) was administered via intravitreal injection. PLR was recorded in response to light stimuli of various intensities. DAMGO eliminated PLR evoked by light with intensities below melanopsin activation threshold but not that evoked by bright blue irradiance that activated melanopsin signaling, although in the latter case, DAMGO markedly slowed pupil constriction. CTAP or genetic ablation of MORs in ipRGCs slightly enhanced dim-light-evoked PLR but not that evoked by a bright blue stimulus. Our results suggest that endogenous opioid signaling in the retina contributes to the regulation of PLR. The slowing of bright light-evoked PLR by DAMGO is consistent with the observation that systemically applied opioids accumulate in the vitreous and that patients receiving chronic opioid treatment have slow PLR.

Cost-effectiveness analysis of reflex testing for Lynch syndrome in women with endometrial cancer in the UK setting.

BACKGROUND: Lynch syndrome is a hereditary cancer syndrome caused by constitutional pathogenic variants in the DNA mismatch repair (MMR) system, leading to increased risk of colorectal, endometrial and other cancers. The study aimed to identify the incremental costs and consequences of strategies to identify Lynch syndrome in women with endometrial cancer. METHODS: A decision-analytic model was developed to evaluate the relative cost-effectiveness of reflex testing strategies for identifying Lynch syndrome in women with endometrial cancer taking the NHS perspective and a lifetime horizon. Model input parameters were sourced from various published sources. Consequences were measured using quality-adjusted life years (QALYs). A cost-effectiveness threshold of £20 000/QALY was used. RESULTS: Reflex testing for Lynch syndrome using MMR immunohistochemistry and MLH1 methylation testing was cost-effective versus no testing, costing £14 200 per QALY gained. There was uncertainty due to parameter imprecision, with an estimated 42% chance this strategy is not cost-effective compared with no testing. Age had a significant impact on cost-effectiveness, with testing not predicted to be cost-effective in patients aged 65 years and over. CONCLUSIONS: Testing for Lynch syndrome in younger women with endometrial cancer using MMR immunohistochemistry and MLH1 methylation testing may be cost-effective. Age cut-offs may be controversial and adversely affect implementation.

A genome-wide association study on photic sneeze reflex in the Chinese population.

Photic sneeze reflex (PSR) is an interesting but yet mysterious phenotype featured by individuals' response of sneezing in exposure to bright light. To uncover the underlying genetic markers (single nucleotide polymorphisms, SNPs), a genome-wide association study (GWAS) was conducted exclusively in a Chinese population of 3417 individuals (PSR prevalence at 25.6%), and reproducibly identified both a replicative rs10427255 on 2q22.3 and a novel locus of rs1032507 on 3p12.1 in various effect models (additive, as well as dominant and recessive). Minor alleles respectively contributed to increased or reduced risk for PSR with odds ratio (95% confidence interval) at 1.68 ([1.50, 1.88]) for rs10427255 and 0.65 ([0.58, 0.72]) for rs1032507. The two independent SNPs were intergenic, and collectively enhanced PSR classification by lifting the area-under-curve value in ROC curve to 0.657. Together with previous GWAS in other populations, the result substantiated the polygenic and non-ethnicity-specific nature behind the PSR phenotype.

Insights into the roles of CMK-1 and OGT-1 in interstimulus interval-dependent habituation in Caenorhabditis elegans.

Habituation is a ubiquitous form of non-associative learning observed as a decrement in responding to repeated stimulation that cannot be explained by sensory adaptation or motor fatigue. One of the defining characteristics of habituation is its sensitivity to the rate at which training stimuli are presented-animals habituate faster in response to more rapid stimulation. The molecular mechanisms underlying this interstimulus interval (ISI)-dependent characteristic of habituation remain unknown. In this article, we use behavioural neurogenetic and bioinformatic analyses in the nematode Caenorhabiditis elegans to identify the first molecules that modulate habituation in an ISI-dependent manner. We show that the Caenorhabditis elegans orthologues of Ca2+/calmodulin-dependent kinases CaMK1/4, CMK-1 and O-linked N-acetylglucosamine (O-GlcNAc) transferase, OGT-1, both function in primary sensory neurons to inhibit habituation at short ISIs and promote it at long ISIs. In addition, both cmk-1 and ogt-1 mutants display a rare mechanosensory hyper-responsive phenotype (i.e. larger mechanosensory responses than wild-type). Overall, our work identifies two conserved genes that function in sensory neurons to modulate habituation in an ISI-dependent manner, providing the first insights into the molecular mechanisms underlying the universally observed phenomenon that habituation has different properties when stimuli are delivered at different rates.

Morpholino-Mediated Exon Inclusion for SMA.

The application of antisense oligonucleotides (AONs) to modify pre-messenger RNA splicing has great potential for treating genetic diseases. The strategies used to redirect splicing for therapeutic purpose involve the use of AONs complementary to splice motifs, enhancer or silencer sequences. AONs to block intronic splicing silencer motifs can efficiently augment exon 7 inclusion in survival motor neuron 2 (SMN2) gene and have demonstrated robust therapeutic effects in both preclinical studies and clinical trials in spinal muscular atrophy (SMA), which has led to a recently approved drug. AONs with phosphorodiamidate morpholino oligomer (PMO) backbone have shown target engagement with restoration of the defective protein in Duchenne muscular dystrophy (DMD) and their safety profile lead to a recent conditional approval for one DMD PMO drug. PMO AONs are also effective in correcting SMN2 exon 7 splicing and rescuing SMA transgenic mice. Here we provide the details of methods that our lab has used to evaluate PMO-mediated SMN2 exon 7 inclusion in the in vivo studies conducted in SMA transgenic mice. The methods comprise mouse experiment procedures, assessment of PMOs on exon 7 inclusion at RNA levels by reverse transcription (RT-) PCR and quantitative real-time PCR. In addition, we present methodology for protein quantification using western blot in mouse tissues, on neuropathology assessment of skeletal muscle (muscle pathology and neuromuscular junction staining) as well as behaviour test in the SMA mice (righting reflex).

Protective effects of Withania somnifera extract in SOD1G93A mouse model of amyotrophic lateral sclerosis.

Withania somnifera (WS; commonly known as Ashwagandha or Indian ginseng) is a medicinal plant whose extracts have been in use for centuries in various regions of the world as a rejuvenator. There is now a growing body of evidence documenting neuroprotective functions of the plant extracts or its purified compounds in several models of neurodegenerative diseases including amyotrophic lateral sclerosis (ALS). Based on the extract's beneficial effect in a mouse model of ALS with TDP-43 proteinopathy, the current study was designed to test its efficacy in another model of familial ALS. Our results show that administration of WS extracts by gavage to mice expressing G93A mutant form of superoxide dismutase (SOD1) resulted in increased longevity, improved motor performance and increased number of motor neurons in lumbar spinal cord. The WS treatment caused substantial reduction in levels of misfolded SOD1whereas it enhanced expression of cellular chaperons in spinal cord of SOD1G93A mice. WS markedly reduced glial activation and prevented phosphorylation of nuclear factor kappaB (NF-κB). The overall immunomodulatory effect of WS was further evidenced by changes in expression of multiple cytokines/chemokines. WS also served as an autophagy inducer which may be beneficial at early stages of the disease. These results suggest that WS extracts might constitute promising therapeutics for treatment of ALS with involvement of misfolded SOD1.

A genome-wide association study on photic sneeze syndrome in a Japanese population.

Photic sneeze syndrome (PSS) is characterized by a tendency to sneeze when the eye is exposed to bright light. Recent genome-wide association studies (GWASs) have identified single-nucleotide polymorphisms (SNPs) associated with PSS in Caucasian populations. We performed a GWAS on PSS in Japanese individuals who responded to a web-based survey and provided saliva samples. After quality control, genotype data of 210,086 SNPs in 11,409 individuals were analyzed. The overall prevalence of PSS was 3.2%. Consistent with previous reports, SNPs at 3p12.1 were associated with PSS at genome-wide significance (p < 5.0 × 10-8). Furthermore, two novel loci at 9q34.2 and 4q35.2 reached suggestive significance (p < 5.0 × 10-6). Our data also provided evidence supporting the two additional SNPs on 2q22.3 and 9q33.2 reportedly associated with PSS. Our study reproduced previous findings in Caucasian populations and further suggested novel PSS loci in the Japanese population.

Motor function deficits in the 12 month-old female 5xFAD mouse model of Alzheimer's disease.

Motor problems occur early in some patients with Alzheimer's disease (AD) and as the disease progresses many patients develop motor dysfunction. Motor dysfunction has been reported in some mouse models of AD, including the 5xFAD mouse, thus this model may be particularly useful for studying motor dysfunction in AD. In order to determine the extent of motor dysfunction in these mice, we tested 11-13 month old female 5xFAD and wildtype (WT) control mice in a battery of motor behaviour tasks. The 5xFAD mice showed hind limb clasping, weighed less and had slower righting reflexes than WT mice. In the open field, the 5xFAD mice travelled a shorter distance than the WT mice, spent less time moving and had a slower movement speed. The 5xFAD mice fell faster than the WT mice from the balance beam, wire suspension, grid suspension and rotarod tasks, indicating dysfunctions in balance, grip strength, motor co-ordination and motor learning. The 5xFAD mice had a short, shuffling gait with a shorter stride length than WT mice and had a slower swim speed. The 5xFAD mice also failed to show an acoustic startle response, likely due to motor dysfunction and previously reported hearing impairment. The 5xFAD mice did not show deficits in the ability of peripheral motor nerves to drive muscle output, suggesting that motor impairments are not due to dysfunction in peripheral motor nerves. These results indicate that the aged 5xFAD mice are deficient in numerous motor behaviours, and suggest that these mice may prove to be a good model for studying the mechanisms of motor dysfunction in AD, and motor behaviour might prove useful for assessing the efficacy of AD therapeutics. Motor dysfunction in 5xFAD mice must also be considered in behavioural tests of sensory and cognitive function so that performance is not confounded by impaired locomotor or swimming behaviour.

Increased ethanol drinking in "humanized" mice expressing the mu opioid receptor A118G polymorphism are mediated through sex-specific mechanisms.

The A118G single nucleotide polymorphism (SNP) of the mu-opioid receptor gene (Oprm1) has been implicated in mediating the rewarding effects of alcohol. Clinical and preclinical studies suggest that the G allele may confer a genetic vulnerability to alcohol dependence, though it remains unknown whether these effects are sex-specific. We used male and female mice homozygous for the "humanized" 118AA or 118GG alleles to determine whether the A118G SNP potentiates ethanol consumption in a sex-specific manner in both the two-bottle choice and drinking-in-the-dark (DID) paradigms. Mice were also assessed for differences in naltrexone sensitivity, ethanol reward assessed via conditioned place preference (CPP), and sensitivity to the sedative/ataxic effects of ethanol using the rota-rod and loss of righting reflex (LORR) assays. We found that male and female 118GG mice drank significantly more ethanol than 118AA littermates using a continuous access, two-bottle choice paradigm. In the limited-access DID drinking model, (i) female (but not male) 118GG mice consumed more ethanol than 118AA mice and (ii) naltrexone pretreatment was equally efficacious at attenuating ethanol intake in both 118AA and 118GG female mice while having no effect in males. Male and female 118GG and female 118AA mice developed a robust conditioned place preference (CPP) for ethanol. Female 118GG mice displayed less sensitivity to the sedative/ataxic effects of ethanol compared to female 118AA mice on both the rota-rod and the LORR assays while male mice did not differ in their responses on either assay. Our findings suggest that increased ethanol consumption in male 118GG mice may be due to increased ethanol reward, while increased drinking in female 118GG mice might be due to decreased sensitivity to the sedative/ataxic effects of ethanol. Collectively, these data might be used to help identify sex-specific pharmacotherapies to combat alcohol use disorders.

RORß Spinal Interneurons Gate Sensory Transmission during Locomotion to Secure a Fluid Walking Gait.

Animals depend on sensory feedback from mechanosensory afferents for the dynamic control of movement. This sensory feedback needs to be selectively modulated in a task- and context-dependent manner. Here, we show that inhibitory interneurons (INs) expressing the RORß orphan nuclear receptor gate sensory feedback to the spinal motor system during walking and are required for the production of a fluid locomotor rhythm. Genetic manipulations that abrogate inhibitory RORß IN function result in an ataxic gait characterized by exaggerated flexion movements and marked alterations to the step cycle. Inactivation of RORß in inhibitory neurons leads to reduced presynaptic inhibition and changes to sensory-evoked reflexes, arguing that the RORß inhibitory INs function to suppress the sensory transmission pathways that activate flexor motor reflexes and interfere with the ongoing locomotor program. VIDEO ABSTRACT.

A Sensitized IGF1 Treatment Restores Corticospinal Axon-Dependent Functions.

A major hurdle for functional recovery after both spinal cord injury and cortical stroke is the limited regrowth of the axons in the corticospinal tract (CST) that originate in the motor cortex and innervate the spinal cord. Despite recent advances in engaging the intrinsic mechanisms that control CST regrowth, it remains to be tested whether such methods can promote functional recovery in translatable settings. Here we show that post-lesional AAV-assisted co-expression of two soluble proteins, namely insulin-like growth factor 1 (IGF1) and osteopontin (OPN), in cortical neurons leads to robust CST regrowth and the recovery of CST-dependent behavioral performance after both T10 lateral spinal hemisection and a unilateral cortical stroke. In these mice, a compound able to increase axon conduction, 4-aminopyridine-3-methanol, promotes further improvement in CST-dependent behavioral tasks. Thus, our results demonstrate a potentially translatable strategy for restoring cortical dependent function after injury in the adult.

Transient receptor potential melastatin 2 channels (TRPM2) mediate neonatal hypoxic-ischemic brain injury in mice.

Transient receptor potential melastatin 2 (TRPM2), a calcium-permeable non-selective cation channel, is reported to mediate brain damage following ischemic insults in adult mice. However, the role of TRPM2 channels in neonatal hypoxic-ischemic brain injury remains unknown. We hypothesize that TRPM2+/- and TRPM2-/- neonatal mice have reduced hypoxic-ischemic brain injury. To study the effect of TRPM2 on neonatal brain damage, we used 2,3,5-triphenyltetrazolium chloride (TTC) staining to assess the infarct volume and whole brain imaging to assess morphological changes in the brain. In addition, we also evaluated neurobehavioral outcomes for sensorimotor function 7days following hypoxic-ischemic brain injury. We report that the infarct volumes were significantly smaller and behavioral outcomes were improved in both TRPM2+/- and TRPM2-/- mice compared to that of wildtype mice. Next, we found that TRPM2-null mice showed reduced dephosphorylation of GSK-3ß following hypoxic ischemic injury unlike sham mice. TRPM2+/- and TRPM2-/- mice also had reduced activation of astrocytes and microglia in ipsilateral hemispheres, compared to wildtype mice. These findings suggest that TRPM2 channels play an essential role in mediating hypoxic-ischemic brain injury in neonatal mice. Genetically eliminating TRPM2 channels can provide neuroprotection against hypoxic-ischemic brain injury and this effect is elicited in part through regulation of GSK-3ß.

Gonadotropin-releasing hormone receptor (Gnrhr) gene knock out: Normal growth and development of sensory, motor and spatial orientation behavior but altered metabolism in neonatal and prepubertal mice.

Gonadotropin-releasing hormone (GnRH) is important in the control of reproduction, but its actions in non-reproductive processes are less well known. In this study we examined the effect of disrupting the GnRH receptor in mice to determine if growth, metabolism or behaviors that are not associated with reproduction were affected. To minimize the effects of other hormones such as FSH, LH and sex steroids, the neonatal-prepubertal period of 2 to 28 days of age was selected. The study shows that regardless of sex or phenotype in the Gnrhr gene knockout line, there was no significant difference in the daily development of motor control, sensory detection or spatial orientation among the wildtype, heterozygous or null mice. This included a series of behavioral tests for touch, vision, hearing, spatial orientation, locomotory behavior and muscle strength. Neither the daily body weight nor the final weight on day 28 of the kidney, liver and thymus relative to body weight varied significantly in any group. However by day 28, metabolic changes in the GnRH null females compared with wildtype females showed a significant reduction in inguinal fat pad weight normalized to body weight; this was accompanied by an increase in glucose compared with wildtype females shown by Student-Newman-Keuls Multiple Comparison test and Student's unpaired t tests. Our studies show that the GnRH-GnRHR system is not essential for growth or motor/sensory/orientation behavior during the first month of life prior to puberty onset. The lack of the GnRH-GnRHR axis, however, did affect females resulting in reduced subcutaneous inguinal fat pad weight and increased glucose with possible insulin resistance; the loss of the normal rise of estradiol at postnatal days 15-28 may account for the altered metabolism in the prepubertal female pups.

Piezo2 senses airway stretch and mediates lung inflation-induced apnoea.

Respiratory dysfunction is a notorious cause of perinatal mortality in infants and sleep apnoea in adults, but the mechanisms of respiratory control are not clearly understood. Mechanical signals transduced by airway-innervating sensory neurons control respiration; however, the physiological significance and molecular mechanisms of these signals remain obscured. Here we show that global and sensory neuron-specific ablation of the mechanically activated ion channel Piezo2 causes respiratory distress and death in newborn mice. Optogenetic activation of Piezo2+ vagal sensory neurons causes apnoea in adult mice. Moreover, induced ablation of Piezo2 in sensory neurons of adult mice causes decreased neuronal responses to lung inflation, an impaired Hering-Breuer mechanoreflex, and increased tidal volume under normal conditions. These phenotypes are reproduced in mice lacking Piezo2 in the nodose ganglion. Our data suggest that Piezo2 is an airway stretch sensor and that Piezo2-mediated mechanotransduction within various airway-innervating sensory neurons is critical for establishing efficient respiration at birth and maintaining normal breathing in adults.

Somatosensory reflex seizures in a child with epilepsy related to novel SCN1A mutation.

INTRODUCTION: Mutations in SCN1A have been reported in patients with different types of epilepsy, including generalized epilepsy with febrile seizures plus, severe myoclonic epilepsy in infancy, malignant migrating partial seizures in infancy, and other infantile epileptic encephalopathies. CASE REPORT: We report a case of a 10-month-old girl presented with reflex epileptic seizures provoked by somatosensory stimuli with a novel de novo mutation of SCN1A gene. She was observed to have seizures with eye deviation, unresponsiveness provoked by somatosensory stimuli of the face. Video-electroencephalography (EEG) revealed generalized spike-and-wave patterns. She experienced one or two focal clonic seizures per month over the 6 months while taking valproate and carbamazepine. At 22 months old, she was hospitalized with an episode of generalized tonic clonic febrile status epilepticus lasting for 45 min. Interictal sleep video-EEG showed sharp-and-slow wave discharges in the left occipital lobe with normal background activity. We found a de novo heterozygote mutation in SCN1A gene, c.1337A>C (p. Q422P). CONCLUSION: To our knowledge, this mutation has not been previously described in the SCN1A gene and this is the first report of epilepsy related to SCN1A mutation as a presenting with reflex epilepsy of somatosensory stimuli. This case report contributes to an expanding clinical spectrum of patients with SCN1A mutations.

Genetic associations with reflexive visual attention in infancy and childhood.

This study elucidates genetic influences on reflexive (as opposed to sustained) attention in children (aged 9-16 years; N = 332) who previously participated as infants in visual attention studies using orienting to a moving bar (Dannemiller, 2004). We investigated genetic associations with reflexive attention measures in infancy and childhood in the same group of children. The genetic markers (single nucleotide polymorphisms and variable number tandem repeats on the genes APOE, BDNF, CHRNA4, COMT, DRD4, HTR4, IGF2, MAOA, SLC5A7, SLC6A3, and SNAP25) are related to brain development and/or to the availability of neurotransmitters such as acetylcholine, dopamine, or serotonin. This study shows that typically developing children have differences in reflexive attention associated with their genes, as we found in adults (Lundwall, Guo & Dannemiller, 2012). This effort to extend our previous findings to outcomes in infancy and childhood was necessary because genetic influence may differ over the course of development. Although two of the genes that were tested in our adult study (Lundwall et al., 2012) were significant in either our infant study (SLC6A3) or child study (DRD4), the specific markers tested differed. Performance on the infant task was associated with SLC6A3. In addition, several genetic associations with an analogous child task occurred with markers on CHRNA4, COMT, and DRD4. Interestingly, the child version of the task involved an interaction such that which genotype group performed poorer on the child task depended on whether we were examining the higher or lower infant scoring group. These findings are discussed in terms of genetic influences on reflexive attention in infancy and childhood.

Deficit in emotional learning in neurotrimin knockout mice.

Neurotrimin (Ntm) belongs to the IgLON family of cell adhesion molecules with Lsamp, Obcam and kilon that regulate the outgrowth of neurites mostly by forming heterodimers. IgLONs have been associated with psychiatric disorders, intelligence, body weight, heart disease and tumours. This study provides an initial behavioural and pharmacological characterization of the phenotype of Ntm-deficient mice. We expected to see at least some overlap with the phenotype of Lsamp-deficient mice as Ntm and Lsamp are the main interaction partners in the IgLON family and are colocalized in some brain regions. However, Ntm-deficient mice displayed none of the deviations in behaviour that we have previously shown in Lsamp-deficient mice, but differently from Lsamp-deficient mice, had a deficit in emotional learning in the active avoidance task. The only overlap was decreased sensitivity to the locomotor stimulating effect of amphetamine in both knockout models. Thus, despite being interaction partners, on the behavioural level Lsamp seems to play a much more central role than Ntm and the roles of these two proteins seem to be complementary rather than overlapping.

¿Qué se entiende por extremidades y miembros?, ¿existe el reflejo axonal? / What do we understand by extremities and limbs? Does the axonal reflex exist?

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¿Qué se entiende por extremidades y miembros?, ¿existe el reflejo axonal?. Réplica / What do we understand by extremities and limbs? Does the axonal reflex exist?. Reply

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