Interictal epileptiform discharges impair word recall in multiple brain areas.

OBJECTIVES: Interictal epileptiform discharges (IEDs) have been linked to memory impairment, but the spatial and temporal dynamics of this relationship remain elusive. In the present study, we aim to systematically characterize the brain areas and times at which IEDs affect memory. METHODS: Eighty epilepsy patients participated in a delayed free recall task while undergoing intracranial electroencephalography (EEG) monitoring. We analyzed the locations and timing of IEDs relative to the behavioral data in order to measure their effects on memory. RESULTS: Overall IED rates did not correlate with task performance across subjects (r = 0.03, p = 0.8). However, at a finer temporal scale, within-subject memory was negatively affected by IEDs during the encoding and recall periods of the task but not during the rest and distractor periods (p < 0.01, p < 0.001, p = 0.3, and p = 0.8, respectively). The effects of IEDs during encoding and recall were stronger in the left hemisphere than in the right (p < 0.05). Of six brain areas analyzed, IEDs in the inferior-temporal, medial-temporal, and parietal areas significantly affected memory (false discovery rate < 0.05). SIGNIFICANCE: These findings reveal a network of brain areas sensitive to IEDs with key nodes in temporal as well as parietal lobes. They also demonstrate the time-dependent effects of IEDs in this network on memory.

Analytical validation of a semi-automated methodology for quantitative measurement of SARS-CoV-2 RNA in wastewater collected in northern New England.

Wastewater-based epidemiology (WBE) can be used to monitor the community presence of infectious disease pathogens of public health concern such as the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Viral nucleic acid has been detected in the stool of SARS-CoV-2-infected individuals. Asymptomatic SARS-CoV-2 infections make community monitoring difficult without extensive and continuous population screening. In this study, we validated a procedure that includes manual pre-processing, automated SARS-CoV-2 RNA extraction and detection workflows using both reverse-transcriptase quantitative polymerase chain reaction (RT-qPCR) and reverse transcriptase droplet digital PCR (RT-ddPCR). Genomic RNA and calibration materials were used to create known concentrations of viral material to determine the linearity, accuracy, and precision of the wastewater extraction and SARS-CoV-2 RNA detection. Both RT-qPCR and RT-ddPCR perform similarly in all the validation experiments, with a limit of detection of 50 copies/mL. A wastewater sample from a care facility with a known outbreak was assessed for viral content in replicate, and we showed consistent results across both assays. Finally, in a 2-week survey of two New Hampshire cities, we assessed the suitability of our methods for daily surveillance. This paper describes the technical validation of a molecular assay that can be used for long-term monitoring of SARS-CoV-2 in wastewater as a potential tool for community surveillance to assist with public health efforts.IMPORTANCEThis paper describes the technical validation of a molecular assay that can be used for the long-term monitoring of SARS-CoV-2 in wastewater as a potential tool for community surveillance to assist with public health efforts.

Chronic morphine increases Fos-positive neurons after concurrent cornea and tail stimulation.

OBJECTIVE: The aim of the present study was to examine the effect of chronic morphine exposure on diffuse noxious inhibitory controls in a large population of neurons throughout the medullary dorsal horn, as assessed using immunocytochemistry for c-Fos protein. BACKGROUND: Overuse of medications, including the opioids, to treat migraine headache can lead to progressively more frequent headaches. In addition, chronic daily headache sufferers and chronic opioid users both lack the inhibition of pain produced by noxious stimulation of a distal body region, often referred to as diffuse noxious inhibitory controls. METHODS: In urethane anesthetized rats, Fos-positive neurons were quantified in chronic morphine and vehicle-treated animals following 52°C noxious thermal stimulation of the cornea with and without the application of a spatially remote noxious stimulus (placement of the tail in 55°C water). RESULTS: When compared to chronic morphine-treated animals that did not receive the spatially remote noxious stimulus, chronic morphine-treated animals given corneal stimulation along with the spatially remote noxious stimulus demonstrated a 163% increase (P < .05) in the number of Fos-positive neurons in the superficial laminae of the medullary dorsal horn and a 682% increase (P < .01) in deep laminae that was restricted to the side ipsilateral to the applied stimulus. In contrast, no significant difference was found in Fos-like immunoreactivity in vehicle-treated animals given concurrent cornea and tail stimulation or only cornea stimulation in either superficial or deep laminae. CONCLUSIONS: It is proposed that an increase in descending facilitation and subsequent loss of diffuse noxious inhibitory controls contributes to the development of medication overuse headache.

Wastewater-Based SARS-CoV-2 Surveillance in Northern New England.

SARS-CoV-2 viral RNA is shed in the stool of 55-70% of infected individuals and can be detected in community wastewater up to 7 days before people present with COVID-19 symptoms. The detection of SARS-CoV-2 RNA in wastewater may serve as a lead indicator of increased community transmission. Here, we monitored viral concentrations in samples collected from nine municipal wastewater facilities in New Hampshire (NH) and Vermont (VT).Twenty-four-h composite primary influent wastewater samples were collected from nine municipal wastewater treatment facilities twice per week for 5 months (late September 2020 to early February 2021). Wastewater was centrifuged for 30 min at 4600 × g, then the supernatant was frozen until further analysis. Once thawed, samples were concentrated, extracted, and tested for SARS-CoV-2 RNA using reverse transcriptase-quantitative PCR (RT-qPCR) and reverse transcriptase-droplet digital PCR (RT-ddPCR) detection methods. Active case counts for each municipality were tracked from the NH and VT state COVID-19 dashboards. We received a total of 283 wastewater samples from all sites during the study period. Viral RNA was detected in 175/283 (61.8%) samples using RT-qPCR and in 195/283 (68.9%) samples using RT-ddPCR. All nine sites showed positivity in the wastewater, with 8/9 (88.8%) sites having over 50% of their samples test positive over the course of the study. Larger municipalities, such as Nashua, Concord, and Lebanon, NH, showed that SARS-CoV-2 positivity in the wastewater can precede spikes in active COVID-19 case counts by as much as 7 days. Smaller municipalities, such as Woodsville, NH and Hartford, VT, showed sporadic SARS-COV-2 detection and did not always precede a rise in active case counts. We detected SARS-CoV-2 RNA in samples from all 9 municipalities tested, including cities and small towns within this region, and showed wastewater positivity as an early indicator of active case count increases in some regions. Some of the smaller rural municipalities with low case counts may require more frequent sampling to detect SARS-CoV-2 in wastewater before a case surge. With timely collection and analysis of wastewater samples, a community could potentially respond to results by increasing public health initiatives, such as tightening mask mandates and banning large indoor gatherings, to mitigate community transmission of SARS-CoV-2. IMPORTANCE Despite vaccination efforts, the delta and omicron variants of SARS-CoV-2 have caused global surges of COVID-19. As the COVID-19 pandemic continues, it is important to find new ways of tracking early signs of SARS-CoV-2 outbreaks. The manuscript outlines how to collect wastewater from treatment facilities, concentrate the virus in a dilute wastewater sample, and detect it using two sensitive PCR-based methods. It also describes important trends in SARS-CoV-2 concentration in wastewater of a rural region of the United States from Fall 2020 - Winter 2021 and demonstrates the utility of wastewater monitoring as a leading indicator of active SARS-CoV-2 cases. Monitoring changes in concentration of SARS-CoV-2 virus in wastewater may offer an early indicator of increased case counts and enable appropriate public health actions to be taken.

T2 relaxation time post febrile status epilepticus predicts cognitive outcome.

Evidence from animal models and patient data indicates that febrile status epilepticus (FSE) in early development can result in permanently diminished cognitive abilities. To understand the variability in cognitive outcome following FSE, we used MRI to measure dynamic brain metabolic responses to the induction of FSE in juvenile rats. We then compared these measurements to the ability to learn an active avoidance spatial task weeks later. T2 relaxation times were significantly lower in FSE rats that were task learners in comparison to FSE non-learners. While T2 time in whole brain held the greatest predictive power, T2 in hippocampus and basolateral amygdala were also excellent predictors. These signal differences in response to FSE indicate that rats that fail to meet metabolic and oxygen demand are more likely to develop spatial cognition deficits. Place cells from FSE non-learners had significantly larger firing fields and higher in-field firing rate than FSE learners and control animals and imply increased excitability in the pyramidal cells of FSE non-learners. These findings suggest a mechanistic cause for the spatial memory deficits in active avoidance and are relevant to other acute neurological insults in early development where cognitive outcome is a concern.

Short duration waveforms recorded extracellularly from freely moving rats are representative of axonal activity.

While extracellular somatic action potentials from freely moving rats have been well characterized, axonal activity has not. We report direct extracellular tetrode recordings of putative axons whose principal feature is a short duration waveform (SDW) with an average peak-trough length less than 179 µs. While SDW recordings using tetrodes have previously been treated as questionable or classified as cells, we hypothesize that they are representative of axonal activity. These waveforms have significantly shorter duration than somatic action potentials, are triphasic and are therefore similar to classic descriptions of microelectrode recordings in white matter and of in vitro action potential propagation along axons. We describe SDWs recorded from pure white-matter tracts including the alveus and corpus callosum. Recordings of several SDWs in the alveus exhibit grid-like firing patterns suggesting these axons carry spatial information from entorhinal cortical neurons. Finally, we locally injected the GABAA agonist Muscimol into layer CA1 of the hippocampus while simultaneously recording somatic activity and SDWs on the same tetrodes. The persistent activity of SDWs during Muscimol inactivation of somatic action potentials indicates that SDWs are representative of action potential propagation along axons projecting from more distal somata. This characterization is important as it illustrates the dangers of exclusively using spike duration as the sole determinant of unit type, particularly in the case of interneurons whose peak-trough times overlap with SDWs. It may also allow future studies to explore how axonal projections from disparate brain regions integrate spatial information in the hippocampus, and provide a basis for studying the effects of pharmaceutical agents on signal transmission in axons, and ultimately to aid in defining the potential role of axons in cognition.

Enrichment and training improve cognition in rats with cortical malformations.

Children with malformations of cortical development (MCD) frequently have associated cognitive impairments which reduce quality of life. We hypothesized that cognitive deficits associated with MCD can be improved with environmental manipulation or additional training. The E17 methylazoxymethanol acetate (MAM) exposure model bears many anatomical hallmarks seen in human MCDs as well as similar behavioral and cognitive deficits. We divided control and MAM exposed Sprague-Dawley rats into enriched and non-enriched groups and tested performance in the Morris water maze. Another group similarly divided underwent sociability testing and also underwent Magnetic Resonance Imaging (MRI) scans pre and post enrichment. A third group of control and MAM rats without enrichment were trained until they reached criterion on the place avoidance task. MAM rats had impaired performance on spatial tasks and enrichment improved performance of both control and MAM animals. Although MAM rats did not have a deficit in sociability they showed similar improvement with enrichment as controls. MRI revealed a whole brain volume decrease with MAM exposure, and an increase in both MAM and control enriched volumes in comparison to non-enriched animals. In the place avoidance task, MAM rats required approximately 3 times as long to reach criterion as control animals, but with additional training were able to reach control performance. Environmental manipulation and additional training can improve cognition in a rodent MCD model. We therefore suggest that patients with MCD may benefit from appropriate alterations in educational strategies, social interaction and environment. These factors should be considered in therapeutic strategies.

Menthol activation of corneal cool cells induces TRPM8-mediated lacrimation but not nociceptive responses in rodents.

PURPOSE: Stimulation to the cornea via noxious chemical and mechanical means evokes tearing, blinking, and pain. In contrast, mild cooling of the ocular surface has been reported to increase lacrimation via activation of corneal cool primary afferent neurons. The purpose of our study was to determine whether menthol induces corneal cool cell activity and lacrimation via the transient receptor potential melastatin-8 (TRPM8) channel without evoking nociceptive responses. METHODS: Tear measurements were made using a cotton thread in TRPM8 wild type and knockout mice after application of menthol (0.05-50 mM) to the cornea. In additional studies, nocifensive responses (eye swiping and lid closure) were quantified following cornea menthol application. Trigeminal ganglion electrophysiologic single unit recordings were performed in rats to determine the effect of low and high concentrations of menthol on corneal cool cells. RESULTS: At low concentrations, menthol increased tear production in TRPM8 wild type and heterozygous animals, but had no effect in TRPM8 knockout mice, while nocifensive responses remained unaffected. At the highest concentration, menthol (50 mM) increased tearing and nocifensive responses in TRPM8 wild type and knockout animals. A low concentration of menthol (0.1 mM) increased cool cell activity, yet a high concentration of menthol (50 mM) had no effect. CONCLUSIONS: These studies indicated that low concentrations of menthol can increase lacrimation via TRPM8 channels without evoking nocifensive behaviors. At high concentrations, menthol can induce lacrimation and nocifensive behaviors in a TRPM8 independent mechanism. The increase in lacrimation is likely due to an increase in cool cell activity.