2-AG-Mediated Control of GABAergic Signaling Is Impaired in a Model of Epilepsy.

Repeated seizures result in a persistent maladaptation of endocannabinoid (eCB) signaling, mediated part by anandamide signaling deficiency in the basolateral amygdala (BLA) that manifests as aberrant synaptic function and altered emotional behavior. Here, we determined the effect of repeated seizures (kindling) on 2-arachidonoylglycerol (2-AG) signaling on GABA transmission by directly measuring tonic and phasic eCB-mediated retrograde signaling in an in vitro BLA slice preparation from male rats. We report that both activity-dependent and muscarinic acetylcholine receptor (mAChR)-mediated depression of GABA synaptic transmission was reduced following repeated seizure activity. These effects were recapitulated in sham rats by preincubating slices with the 2-AG synthesizing enzyme inhibitor DO34. Conversely, preincubating slices with the 2-AG degrading enzyme inhibitor KML29 rescued activity-dependent 2-AG signaling, but not mAChR-mediated synaptic depression, over GABA transmission in kindled rats. These effects were not attributable to a change in cannabinoid type 1 (CB1) receptor sensitivity or altered 2-AG tonic signaling since the application of the highly selective CB1 receptor agonist CP55,940 provoked a similar reduction in GABA synaptic activity in both sham and kindled rats, while no effect of either DO34 or of the CB1 inverse agonist AM251 was observed on frequency and amplitude of spontaneous IPSCs in either sham or kindled rats. Collectively, these data provide evidence that repeated amygdala seizures persistently alter phasic 2-AG-mediated retrograde signaling at BLA GABAergic synapses, probably by impairing stimulus-dependent 2-AG synthesis/release, which contributes to the enduring aberrant synaptic plasticity associated with seizure activity.SIGNIFICANCE STATEMENT The plastic reorganization of endocannabinoid (eCB) signaling after seizures and during epileptogenesis may contribute to the negative neurobiological consequences associated with seizure activity. Therefore, a deeper understanding of the molecular basis underlying the pathologic long-term eCB signaling remodeling following seizure activity will be crucial to the development of novel therapies for epilepsy that not only target seizure activity, but, most importantly, the epileptogenesis and the comorbid conditions associated with epilepsy.

Complex forelimb movements and cortical topography evoked by intracortical microstimulation in male and female mice.

The motor cortex is crucial for the voluntary control of skilled movement in mammals and is topographically organized into representations of the body (motor maps). Intracortical microstimulation of the motor cortex with long-duration pulse trains (LD-ICMS; ~500 ms) evokes complex movements, occurring in multiple joints or axial muscles, with characteristic movement postures and cortical topography across a variety of mammalian species. Although the laboratory mouse is extensively used in basic and pre-clinical research, high-resolution motor maps elicited with electrical LD-ICMS in both sexes of the adult mouse has yet to be reported. To address this knowledge gap, we performed LD-ICMS of the forelimb motor cortex in both male (n = 10) and naturally cycling female (n = 8) C57/BL6J mice under light ketamine-xylazine anesthesia. Complex and simple movements were evoked from historically defined caudal (CFA) and rostral (RFA) forelimb areas. Four complex forelimb movements were identified consisting of Elevate, Advance, Dig, and Retract postures with characteristic movement sequences and endpoints. Furthermore, evoked complex forelimb movements and cortical topography in mice were organized within the CFA in a unique manner relative to a qualitative comparison with the rat.

Stress-induced modulation of endocannabinoid signaling leads to delayed strengthening of synaptic connectivity in the amygdala.

Even a brief exposure to severe stress strengthens synaptic connectivity days later in the amygdala, a brain area implicated in the affective symptoms of stress-related psychiatric disorders. However, little is known about the synaptic signaling mechanisms during stress that eventually culminate in its delayed impact on the amygdala. Hence, we investigated early stress-induced changes in amygdalar synaptic signaling in order to prevent its delayed effects. Whole-cell recordings in basolateral amygdala (BLA) slices from rats revealed higher frequency of miniature excitatory postsynaptic currents (mEPSCs) immediately after 2-h immobilization stress. This was replicated by inhibition of cannabinoid receptors (CB1R), suggesting a role for endocannabinoid (eCB) signaling. Stress also reduced N-arachidonoylethanolamine (AEA), an endogenous ligand of CB1R. Since stress-induced activation of fatty acid amide hydrolase (FAAH) reduces AEA, we confirmed that oral administration of an FAAH inhibitor during stress prevents the increase in synaptic excitation in the BLA soon after stress. Although stress also caused an immediate reduction in synaptic inhibition, this was not prevented by FAAH inhibition. Strikingly, FAAH inhibition during the traumatic stressor was also effective 10 d later on the delayed manifestation of synaptic strengthening in BLA neurons, preventing both enhanced mEPSC frequency and increased dendritic spine-density. Thus, oral administration of an FAAH inhibitor during a brief stress prevents the early synaptic changes that eventually build up to hyperexcitability in the amygdala. This framework is of therapeutic relevance because of growing interest in targeting eCB signaling to prevent the gradual development of emotional symptoms and underlying amygdalar dysfunction triggered by traumatic stress.

Electrographic seizures and brain hyperoxia may be key etiological factors for postconcussive deficits.

Repetitive mild traumatic brain injuries (RmTBIs) are increasingly recognized to have long-term neurological sequelae in a significant proportion of patients. Individuals that have had RmTBIs exhibit a variety of sensory, cognitive, or behavioral consequences that can negatively impact quality of life. Brain tissue oxygen levels ([Formula: see text]) are normally maintained through exquisite regulation of blood supply to stay within the normoxic zone (18-30 mmHg in the rat hippocampus). However, during neurological events in which brain tissue oxygen levels leave the normoxic zone, neuronal dysfunction and behavioral deficits have been observed, and are frequently related to poorer prognoses. The oxygenation response in the brain after RmTBIs/repeated concussions has been poorly characterized, with most preliminary research limited to the neocortex. Furthermore, the mechanisms by which RmTBIs impact changes to brain oxygenation and vice versa remain to be determined. In the current study, we demonstrate that upon receiving RmTBIs, rats exhibit posttraumatic, electrographic seizures in the hippocampus, without behavioral (clinical) seizures, that are accompanied by a long-lasting period of hyperoxygenation. These electrographic seizures and the ensuing hyperoxic episodes are associated with deficits in working memory and motor coordination that were reversible through attenuation of the posttraumatic and postictal (postseizure) hyperoxia, via administration of a vasoconstricting agent, the calcium channel agonist Bay K8644. We propose that the posttraumatic period characterized by brain oxygenation levels well above the normoxic zone, may be the basis for some of the common symptoms associated with RmTBIs.NEW & NOTEWORTHY We monitor oxygenation and electrographic activity in the hippocampus, immediately before and after mild traumatic brain injury. We demonstrate that as the number of injuries increases from 1 to 3, the proportion of rats that exhibit electrographic seizures and hyperoxia increases. Moreover, the presence of electrographic seizures and hyperoxia are associated with postinjury behavioral impairments, and if the hyperoxia is blocked with Bay K8644, the behavioral deficits are eliminated.

Febrile seizures lead to prolonged epileptiform activity and hyperoxia that when blocked prevents learning deficits.

OBJECTIVE: In adult brain tissue, oxygen levels typically remain in the normoxic zone, but status epilepticus results in hyperoxia, whereas brief self-terminating seizures lead to postictal hypoxia. The dynamic changes in oxygen levels and the underlying mechanisms are unknown in juveniles with febrile seizures. METHODS: Eight-day-old female and male rat pups were implanted with an electrode and oxygen-sensing optode in the hippocampus and then received once daily injections of lipopolysaccharide for 4 days to induce an immune response. Local partial pressure of oxygen (pO2 ) and local field potentials were recorded before, during, and after a heat-induced febrile seizure. Separate groups of pups received injections of vehicle or drugs targeting cyclooxygenase (COX)-1, COX-2, L-type calcium channels (LTCCs), and cannabinoid receptor type 1 (CB1) and transient receptor potential vanilloid-1 (TRPV1) receptors prior to febrile seizure induction to determine pO2 mechanisms. Following febrile seizures, a subset of pups were raised to young adulthood and then tested for learning impairments using the novel object recognition task. RESULTS: Febrile seizures resulted in predictable oxygen dynamics that were related to behavioral seizures and epileptiform activity. During a behavioral seizure, pO2 rapidly increased, rapidly decreased, and then returned to near baseline. When the behavioral seizure terminated, oxygen levels climbed into the hyperoxic zone during a time of prolonged epileptiform activity. When epileptiform activity terminated, oxygen levels slowly returned to baseline. A COX-1 antagonist prevented hyperoxia, whereas a COX-2 antagonist did not. An LTCC antagonist exacerbated hyperoxia. Boosting levels of an endocannabinoid also exacerbated hyperoxia, whereas blocking CB1 receptors and TRPV1 receptors reduced hyperoxia. Inhibiting TRPV1 receptors during a febrile seizure prevented learning deficits in young adult female rats. SIGNIFICANCE: Brain oxygenation during and following a febrile seizure has a distinct pattern and multiple mechanisms. Brain oxygen dynamics may be an important consideration in the development of treatments for febrile seizures.

Anandamide Signaling Augmentation Rescues Amygdala Synaptic Function and Comorbid Emotional Alterations in a Model of Epilepsy.

Epilepsy is often associated with emotional disturbances and the endocannabinoid (eCB) system tunes synaptic transmission in brain regions regulating emotional behavior. Thus, persistent alteration of eCB signaling after repeated seizures may contribute to the development of epilepsy-related emotional disorders. Here we report that repeatedly eliciting seizures (kindling) in the amygdala caused a long-term increase in anxiety and impaired fear memory retention, which was paralleled by an imbalance in GABA/glutamate presynaptic activity and alteration of synaptic plasticity in the basolateral amygdala (BLA), in male rats. Anandamide (AEA) content was downregulated after repeated seizures, and pharmacological enhancement of AEA signaling rescued seizure-induced anxiety by restoring the tonic control of the eCB signaling over glutamatergic transmission. Moreover, AEA signaling augmentation also rescued the seizure-induced alterations of fear memory by restoring the phasic control of eCB signaling over GABAergic activity and plasticity in the BLA. These results indicate that modulation of AEA signaling represents a potential and promising target for the treatment of comorbid emotional dysfunction associated with epilepsy.SIGNIFICANCE STATEMENT Epilepsy is a heterogeneous neurologic disorder commonly associated with comorbid emotional alterations. However, the management of epilepsy is usually restricted to the control of seizures. The endocannabinoid (eCB) system, particularly anandamide (AEA) signaling, controls neuronal excitability and seizure expression and regulates emotional behavior. We found that repeated seizures cause an allostatic maladaptation of AEA signaling in the amygdala that drives emotional alterations. Boosting AEA signaling through inhibition of its degradative enzyme, fatty acid amide hydrolase (FAAH), restored both synaptic and behavioral alterations. FAAH inhibitors dampen seizure activity in animal models and are used in clinical studies to treat the negative consequences associated with stress. Thereby, they are accessible and can be clinically evaluated to treat both seizures and comorbid conditions associated with epilepsy.

Development and plasticity of complex movement representations.

The mammalian motor cortex is topographically organized into representations of discrete body parts (motor maps). Studies in adult rats using long-duration intracortical microstimulation (LD-ICMS) reveal that forelimb motor cortex is functionally organized into several spatially distinct areas encoding complex, multijoint movement sequences: elevate, advance, grasp, and retract. The topographical arrangement of complex movements during development and the influence of skilled learning are unknown. Here, we determined the emergence and topography of complex forelimb movement representations in rats between postnatal days (PND) 13 and 60. We further investigated the expression of the maps for complex movements under conditions of reduced cortical inhibition and whether skilled forelimb motor training could alter their developing topography. We report that simple forelimb movements are first evoked at PND 25 and are confined to the caudal forelimb area (CFA), whereas complex movements first reliably appear at PND 30 and are observed in both the caudal and rostral forelimb areas (RFA). During development, the topography of complex movement representations undergoes reorganization with "grasp" and "elevate" movements predominantly observed in the RFA and all four complex movements observed in CFA. Under reduced cortical inhibition, simple and complex movements were first observed in the CFA on PND 15 and 20, respectively, and the topography is altered relative to a saline control. Further, skilled motor learning was associated with increases in "grasp" and "retract" representations specific to the trained limb. Our results demonstrate that early-life motor experience during development can modify the topography of complex forelimb movement representations.NEW & NOTEWORTHY The motor cortex is topographically organized into maps of different body parts. We used to think that the function of motor cortex was to drive individual muscles, but more recently we have learned that it is also organized to make complex movements. However, the development and plasticity of those complex movements is completely unknown. In this paper, the emergence and topography of complex movement representation, as well as their plasticity during development, is detailed.

The ketogenic diet raises brain oxygen levels, attenuates postictal hypoxia, and protects against learning impairments.

OBJECTIVES: A prolonged vasoconstriction/hypoperfusion/hypoxic event follows self-terminating focal seizures. The ketogenic diet (KD) has demonstrated efficacy as a metabolic treatment for intractable epilepsy and other disorders but its effect on local brain oxygen levels is completely unknown. This study investigated the effects of the KD on tissue oxygenation in the hippocampus before and after electrically elicited (kindled) seizures and whether it could protect against a seizure-induced learning impairment. We also examined the effects of the ketone ß-hydroxybutyrate (BHB) as a potential underlying mechanism. METHODS: Male and female rats were given access to one of three diet protocols 2 weeks prior to the initiation of seizures: KD, caloric restricted standard chow, and ad libitum standard chow. Dorsal hippocampal oxygen levels were measured prior to initiation of diets as well as before and after a 10-day kindling paradigm. Male rats were then tested on a novel object recognition task to assess postictal learning impairments. In a separate cohort, BHB was administered 30 min prior to seizure elicitation to determine whether it influenced oxygen dynamics. RESULTS: The KD increased dorsal hippocampal oxygen levels, ameliorated postictal hypoxia, and prevented postictal learning impairments. Acute BHB administration did not alter oxygen levels before or after seizures. INTERPRETATION: The ketogenic diet raised brain oxygen levels and attenuated severe postictal hypoxia likely through a mechanism independent of ketosis and shows promise as a non-pharmacological treatment to prevent the postictal state.

Reelin Improves Cognition and Extends the Lifespan of Mutant Ndel1 Mice with Postnatal CA1 Hippocampus Deterioration.

The glycoprotein Reelin maintains neuronal positioning and regulates neuronal plasticity in the adult brain. Reelin deficiency has been associated with neurological diseases. We recently showed that Reelin is depleted in mice with a targeted disruption of the Ndel1 gene in forebrain postnatal excitatory neurons (Ndel1 conditional knockout (CKO)). Ndel1 CKO mice exhibit fragmented microtubules in CA1 pyramidal neurons, profound deterioration of the CA1 hippocampus and a shortened lifespan (~10 weeks). Here we report that Ndel1 CKO mice (of both sexes) experience spatial learning and memory deficits that are associated with deregulation of neuronal cell adhesion, plasticity and neurotransmission genes, as assessed by genome-wide transcriptome analysis of the hippocampus. Importantly, a single injection of Reelin protein in the hippocampus of Ndel1 CKO mice improves spatial learning and memory function and this is correlated with reduced intrinsic hyperexcitability of CA1 pyramidal neurons, and normalized gene deregulation in the hippocampus. Strikingly, when treated with Reelin, Ndel1 CKO animals that die from an epileptic phenotype, live twice as long as nontreated, or vehicle-treated CKO animals. Thus, Reelin confers striking beneficial effects in the CA1 hippocampus, and at both behavioral and organismal levels.

Dynamic oxygen changes during status epilepticus and subsequent endogenous kindling.

OBJECTIVE: Brain tissue oxygen (partial oxygen pressure [pO2 ]) levels are tightly regulated to stay within the normoxic zone, with deviations on either side resulting in impaired brain function. Whereas pathological events such as ischemic attacks and brief seizures have previously been shown to result in pO2 levels well below the normoxic zone, oxygen levels during prolonged status epilepticus (SE) and the subsequent endogenous kindling period are unknown. METHODS: We utilized two models of acquired temporal lobe epilepsy in rats: intrahippocampal kainic acid infusion and prolonged perforant pathway stimulation. Local tissue oxygen was measured in the dorsal hippocampus using an optode during and for several weeks following SE. RESULTS: We observed hyperoxia in the hippocampus during induced SE in both models. Following termination of SE, 88% of rats initiated focal self-generated spiking activity in the hippocampus within the first 7 days, which was associated with dynamic oxygen changes. Self-generated and recurring epileptiform activity subsequently organized into higher-frequency bursts that became progressively longer and were ultimately associated with behavioral seizures that became more severe with time and led to postictal hypoxia. SIGNIFICANCE: Induced SE and self-generated recurrent epileptiform activity can have profound and opposing effects on brain tissue oxygenation that may serve as a biomarker for ongoing pathological activity in the brain.

Signs and symptoms of the postictal period in epilepsy: A systematic review and meta-analysis.

OBJECTIVE: The postictal period has many physical, behavioral, and cognitive manifestations associated with it. These signs and symptoms are common, can be quite debilitating, and can have a continued impact long after the seizure has ended. The purpose of this systematic review was to quantify the occurrence of postictal signs and symptoms, along with their frequency and duration in persons with epilepsy. METHODS: Cochrane Database of Systematic Reviews, CINAHL, EMBASE, MEDLINE, PsycINFO, Web of Science, and Scopus were searched from inception to November 29, 2017. The Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) reporting standards were followed. Search terms included subject headings and text words such as convulsion, epilepsy, seizure, postictal, post seizure, seizure recovery, seizure end, Todd's paresis, and Todd's paralysis. Standardized forms were used to collect various study variables. Abstract and full-text review, data abstraction, and quality assessment were all done in duplicate. Study heterogeneity was assessed using the I-squared test, and a random effects model was used to determine estimates. Publication bias was evaluated using funnel plots. RESULTS: From 7811 abstracts reviewed, 78 articles met eligibility criteria, with 31 postictal manifestations (signs and/or symptoms) described and 45 studies included in the meta-analysis. The majority of studies described postictal headaches, migraines, and psychoses, with mean weighted frequency of 33.0% [95% confidence interval (CI) 26.0-40.0], 16.0% [95% CI 10.0-22.0], and 4.0% [95% CI 2.0-5.0], respectively. The mean weighted proportions of manifestations ranged from 0.5% (subacute postictal aggression) to 96.2% (postictal unresponsiveness) with symptom duration usually lasting <24 h but up to 2 months for physical and cognitive/behavioral symptoms respectively. SIGNIFICANCE: Examining data on the various signs and symptoms of the postictal period will have practical applications for physicians by raising their awareness about these manifestations and informing them about the importance of optimizing their prevention and treatment in epilepsy.

Ndel1 and Reelin Maintain Postnatal CA1 Hippocampus Integrity.

UNLABELLED: How the integrity of laminar structures in the postnatal brain is maintained impacts neuronal functions. Ndel1, the mammalian homolog of NuDE from the filamentous fungus Aspergillus nidulans, is an atypical microtubule (MT)-associated protein that was initially investigated in the contexts of neurogenesis and neuronal migration. Constitutive knock-out mice for Ndel1 are embryonic lethal, thereby necessitating the creation a conditional knock-out to probe the roles of Ndel1 in postnatal brains. Here we report that CA1 pyramidal neurons from mice postnatally lacking Ndel1 (Ndel1 conditional knock-out) exhibit fragmented MTs, dendritic/synaptic pathologies, are intrinsically hyperexcitable and undergo dispersion independently of neuronal migration defect. Secondary to the pyramidal cell changes is the decreased inhibitory drive onto pyramidal cells from interneurons. Levels of the glycoprotein Reelin that regulates MTs, neuronal plasticity, and cell compaction are significantly reduced in hippocampus of mutant mice. Strikingly, a single injection of Reelin into the hippocampus of Ndel1 conditional knock-out mice ameliorates ultrastructural, cellular, morphological, and anatomical CA1 defects. Thus, Ndel1 and Reelin contribute to maintain postnatal CA1 integrity. SIGNIFICANCE STATEMENT: The significance of this study rests in the elucidation of a role for Nde1l and Reelin in postnatal CA1 integrity using a new conditional knock-out mouse model for the cytoskeletal protein Ndel1, one that circumvents the defects associated with neuronal migration and embryonic lethality. Our study serves as a basis for understanding the mechanisms underlying postnatal hippocampal maintenance and function, and the significance of decreased levels of Ndel1 and Reelin observed in patients with neurological disorders.

HCN channels segregate stimulation-evoked movement responses in neocortex and allow for coordinated forelimb movements in rodents.

KEY POINTS: The present study tested whether HCN channels contribute to the organization of motor cortex and to skilled motor behaviour during a forelimb reaching task. Experimental reductions in HCN channel signalling increase the representation of complex multiple forelimb movements in motor cortex as assessed by intracortical microstimulation. Global HCN1KO mice exhibit reduced reaching accuracy and atypical movements during a single-pellet reaching task relative to wild-type controls. Acute pharmacological inhibition of HCN channels in forelimb motor cortex decreases reaching accuracy and increases atypical movements during forelimb reaching. ABSTRACT: The mechanisms by which distinct movements of a forelimb are generated from the same area of motor cortex have remained elusive. Here we examined a role for HCN channels, given their ability to alter synaptic integration, in the expression of forelimb movement responses during intracortical microstimulation (ICMS) and movements of the forelimb on a skilled reaching task. We used short-duration high-resolution ICMS to evoke forelimb movements following pharmacological (ZD7288), experimental (electrically induced cortical seizures) or genetic approaches that we confirmed with whole-cell patch clamp to substantially reduce Ih current. We observed significant increases in the number of multiple movement responses evoked at single sites in motor maps to all three experimental manipulations in rats or mice. Global HCN1 knockout mice were less successful and exhibited atypical movements on a skilled-motor learning task relative to wild-type controls. Furthermore, in reaching-proficient rats, reaching accuracy was reduced and forelimb movements were altered during infusion of ZD7288 within motor cortex. Thus, HCN channels play a critical role in the separation of overlapping movement responses and allow for successful reaching behaviours. These data provide a novel mechanism for the encoding of multiple movement responses within shared networks of motor cortex. This mechanism supports a viewpoint of primary motor cortex as a site of dynamic integration for behavioural output.

Postictal hypoperfusion/hypoxia provides the foundation for a unified theory of seizure-induced brain abnormalities and behavioral dysfunction.

A recent article by Farrell et al. characterizes the phenomenon, mechanisms, and treatment of a local and severe hypoperfusion/hypoxia event that occurs in brain regions following a focal seizure. Given the well-established role of cerebral ischemia/hypoxia in brain damage and behavioral dysfunction in other clinical settings (e.g., stroke, cerebral vasospasm), we put forward a new theory: postictal hypoperfusion/hypoxia is responsible for the negative consequences associated with seizures. Fortunately, inhibition of two separate molecular targets, cyclooxygenase-2 (COX-2) and l-type calcium channels, can prevent the expression of postictal hypoperfusion/hypoxia. These inhibitors are important experimental tools used to separate the seizure from the resulting hypoperfusion/hypoxia and can allow researchers to address the contribution of this phenomenon to negative outcomes associated with seizures. Herein we address the implications of this postictal stroke-like event in acute behavioral dysfunction (e.g., Todd's paresis) and sudden unexpected death in epilepsy (SUDEP). Moreover, anatomic alterations such as increased blood-brain barrier permeability, glial activation, central inflammation, and neuronal loss could also be a consequence of repeated hypoperfusion/hypoxic events and, in turn, underlie chronic interictal cognitive and behavioral comorbidities (e.g., memory deficits, anxiety, depression, and psychosis) and exacerbate epileptogenesis. Thus these seemingly disparate and clinically important observations may share a common point of origin: postictal hypoperfusion/hypoxia.

Motivational wheel running reverses cueing behavioural inflexibility in rodents.

Behavioural inflexibility and associated atypical learning behaviours are common clinical manifestations of the autism spectrum disorder (ASD) phenotype. Despite advances in our understanding of ASD, little research has been devoted to experimental interventions that might help to circumvent behavioural inflexibility in ASD. The current paper suggests that motivational locomotion in the form of wheel running can reduce behavioural inflexibility and learning impairments in an ASD rat model, and discusses how the strategy of reward-coupled locomotor activity may lead to clinical interventions for children with ASD.

Motor cortex is functionally organized as a set of spatially distinct representations for complex movements.

There is a long-standing debate regarding the functional organization of motor cortex. Intracortical microstimulation (ICMS) studies have provided two contrasting views depending on the duration of stimulation. In the rat, short-duration ICMS reveals two spatially distributed forelimb movement representations, the rostral forelimb area (RFA) and caudal forelimb area (CFA), eliciting identical movements. In contrast, long-duration ICMS reveals spatially distributed, complex, multijoint movement areas, with grasping found exclusively in the rostral area and reach-shaping movements of the arm located in the caudal area. To provide corroboration for which interpretation is correct, we selectively inactivated the RFA/grasp area during the performance of skilled forelimb behaviors using a reversible cortical cooling deactivation technique. A significant impairment of grasping in the single-pellet retrieval task and manipulations of pasta was observed during cooling deactivation of the RFA/grasp area, but not the CFA/arm area. Our results indicate a movement-based, rather than a muscle-based, functional organization of motor cortex, and provide evidence for a conserved homology of independent grasp and reach circuitry shared between primates and rats.

Experience salience gates endocannabinoid signaling at hypothalamic synapses.

Alterations in synaptic endocannabinoid signaling are a widespread neurobiological consequence of many in vivo experiences, including stress. Here, we report that stressor salience is critical for bidirectionally modifying presynaptic CB-1 receptor (CB1R) function at hypothalamic GABA synapses controlling the neuroendocrine stress axis in male rats. While repetitive, predictable stressor exposure impairs presynaptic CB1R function, these changes are rapidly reversed upon exposure to a high salience experience such as novel stress or by manipulations that enhance neural activity levels in vivo or in vitro. Together these data demonstrate that experience salience, through alterations in afferent synaptic activity, induces rapid changes in endocannabinoid signaling.

Serotonin 1A receptors alter expression of movement representations.

Serotonin has a myriad of central functions involving mood, appetite, sleep, and memory and while its release within the spinal cord is particularly important for generating movement, the corresponding role on cortical movement representations (motor maps) is unknown. Using adult rats we determined that pharmacological depletion of serotonin (5-HT) via intracerebroventricular administration of 5,7 dihydroxytryptamine resulted in altered movements of the forelimb in a skilled reaching task as well as higher movement thresholds and smaller maps derived using high-resolution intracortical microstimulation (ICMS). We ruled out the possibility that reduced spinal cord excitability could account for the serotonin depletion-induced changes as we observed an enhanced Hoffman reflex (H-reflex), indicating a hyperexcitable spinal cord. Motor maps derived in 5-HT1A receptor knock-out mice also showed higher movement thresholds and smaller maps compared with wild-type controls. Direct cortical application of the 5-HT1A/7 agonist 8-OH-DPAT lowered movement thresholds in vivo and increased map size in 5-HT-depleted rats. In rats, electrical stimulation of the dorsal raphe lowered movement thresholds and this effect could be blocked by direct cortical application of the 5-HT1A antagonist WAY-100135, indicating that serotonin is primarily acting through the 5-HT1A receptor. Next we developed a novel in vitro ICMS preparation that allowed us to track layer V pyramidal cell excitability. Bath application of WAY-100135 raised the ICMS current intensity to induce action potential firing whereas the agonist 8-OH-DPAT had the opposite effect. Together our results demonstrate that serotonin, acting through 5-HT1A receptors, plays an excitatory role in forelimb motor map expression.

Caffeine exacerbates seizure-induced death via postictal hypoxia.

Sudden unexpected death in epilepsy (SUDEP) is the leading epilepsy-related cause of premature mortality in people with intractable epilepsy, who are 27 times more likely to die than the general population. Impairment of the central control of breathing following a seizure has been identified as a putative cause of death, but the mechanisms underlying this seizure-induced breathing failure are largely unknown. Our laboratory has advanced a vascular theory of postictal behavioural dysfunction, including SUDEP. We have recently reported that seizure-induced death occurs after seizures invade brainstem breathing centres which then leads to local hypoxia causing breathing failure and death. Here we investigated the effects of caffeine and two adenosine receptors in two models of seizure-induced death. We recorded local oxygen levels in brainstem breathing centres as well as time to cessation of breathing and cardiac activity relative to seizure activity. The administration of the non-selective A1/A2A antagonist caffeine or the selective A1 agonist N6-cyclopentyladenosine reveals a detrimental effect on postictal hypoxia, providing support for caffeine modulating cerebral vasculature leading to brainstem hypoxia and cessation of breathing. Conversely, A2A activation with CGS-21680 was found to increase the lifespan of mice in both our models of seizure-induced death.

Postictal hypoxia involves reactive oxygen species and is ameliorated by chronic mitochondrial uncoupling.

Prolonged severe hypoxia follows brief seizures and represents a mechanism underlying several negative postictal manifestations without interventions. Approximately 50% of the postictal hypoxia phenomenon can be accounted for by arteriole vasoconstriction. What accounts for the rest of the drop in unbound oxygen is unclear. Here, we determined the effect of pharmacological modulation of mitochondrial function on tissue oxygenation in the hippocampus of rats after repeatedly evoked seizures. Rats were treated with mitochondrial uncoupler 2,4 dinitrophenol (DNP) or antioxidants. Oxygen profiles were recorded using a chronically implanted oxygen-sensing probe, before, during, and after seizure induction. Mitochondrial function and redox tone were measured using in vitro mitochondrial assays and immunohistochemistry. Postictal cognitive impairment was assessed using the novel object recognition task. Mild mitochondrial uncoupling by DNP raised hippocampal oxygen tension and ameliorated postictal hypoxia. Chronic DNP also lowered mitochondrial oxygen-derived reactive species and oxidative stress in the hippocampus during postictal hypoxia. Uncoupling the mitochondria exerts therapeutic benefits on postictal cognitive dysfunction. Finally, antioxidants do not affect postictal hypoxia, but protect the brain from associated cognitive deficits. We provided evidence for a metabolic component of the prolonged oxygen deprivation that follow seizures and its pathological sequelae. Furthermore, we identified a molecular underpinning of this metabolic component, which involves excessive oxygen conversion into reactive species. Mild mitochondrial uncoupling may be a potential therapeutic strategy to treat the postictal state where seizure control is absent or poor.