Ampakine pretreatment enables a single brief hypoxic episode to evoke phrenic motor facilitation.

Phrenic long-term facilitation (LTF) is a sustained increase in phrenic motor output occurring after exposure to multiple (but not single) hypoxic episodes. Ampakines are a class of drugs that enhance AMPA receptor function. Ampakines can enhance expression of neuroplasticity, and the phrenic motor system is fundamentally dependent on excitatory glutamatergic currents. Accordingly, we tested the hypothesis that combining ampakine pretreatment with a single brief hypoxic exposure would result in phrenic motor facilitation lasting well beyond the period of hypoxia. Phrenic nerve output was recorded in urethane-anesthetized, ventilated, and vagotomized adult Sprague-Dawley rats. Ampakine CX717 (15 mg/kg iv; n = 8) produced a small increase in phrenic inspiratory burst amplitude and frequency, but values quickly returned to predrug baseline. When CX717 was followed 2 min later by a 5-min exposure to hypoxia (n = 8; PaO2 ~45 mmHg), a persistent increase in phrenic inspiratory burst amplitude (i.e., phrenic motor facilitation) was observed up to 60 min posthypoxia (103 ± 53% increase from baseline). In contrast, when hypoxia was preceded by vehicle injection (10% 2-hydroxypropyl-ß-cyclodextrin; n = 8), inspiratory phrenic bursting was similar to baseline values at 60 min. Additional experiments with another ampakine (CX1739, 15 mg/kg) produced comparable results. We conclude that pairing low-dose ampakine treatment with a single brief hypoxic exposure can evoke sustained phrenic motor facilitation. This targeted approach for enhancing respiratory neuroplasticity may have value in the context of hypoxia-based neurorehabilitation strategies.NEW & NOTEWORTHY A single brief episode of hypoxia (e.g., 3-5 min) does not evoke long-lasting increases in respiratory motor output after the hypoxia is concluded. Ampakines are a class of drugs that enhance AMPA receptor function. We show that pairing low-dose ampakine treatment with a single brief hypoxic exposure can evoke sustained phrenic motor facilitation after the acute hypoxic episode.

When should governments increase the supply of psychiatric beds?

Low numbers of hospital-based psychiatric beds create problems for people with severe mental illness (SMI), when they face extended emergency department (ED) waits, higher thresholds for admission to an acute bed, and short revolving-door stays with high rates of rehospitalisation. Limited access to inpatient treatment has been associated with higher suicide risk, premature mortality, homelessness, violent crime and incarceration. Ultimately, people with SMI can be transinstitutionalised to the criminal justice system. In the USA, for example, prisons have replaced mental hospitals as the largest institutions housing people with SMI. There is no international consensus on the safe minimum numbers of acute, forensic and rehabilitation beds needed to reduce these risks. As a consequence, Organisation for Economic Cooperation and Development (OECD) countries have wide variations in the mix of hospital beds with an average of 71 beds per 100 000 population. Policymakers face difficult choices with few studies to guide decisions on supplying beds. The UK Royal College of Psychiatrists offered a policy framework, which was adapted for Australia. The government of the State of South Australia increased the supplies of crisis, acute and forensic beds to meet a mandatory target to safely reduce mental health boarding in the EDs.

Intraspinal microstimulation and diaphragm activation after cervical spinal cord injury.

Intraspinal microstimulation (ISMS) using implanted electrodes can evoke locomotor movements after spinal cord injury (SCI) but has not been explored in the context of respiratory motor output. An advantage over epidural and direct muscle stimulation is the potential of ISMS to selectively stimulate components of the spinal respiratory network. The present study tested the hypothesis that medullary respiratory activity could be used to trigger midcervical ISMS and diaphragm motor unit activation in rats with cervical SCI. Studies were conducted after acute (hours) and subacute (5-21 days) C2 hemisection (C2Hx) injury in adult rats. Inspiratory bursting in the genioglossus (tongue) muscle was used to trigger a 250-ms train stimulus (100 Hz, 100-200 µA) to the ventral C4 spinal cord, targeting the phrenic motor nucleus. After both acute and subacute injury, genioglossus EMG activity effectively triggered ISMS and activated diaphragm motor units during the inspiratory phase. The ISMS paradigm also evoked short-term potentiation of spontaneous inspiratory activity in the previously paralyzed hemidiaphragm (i.e., bursting persisting beyond the stimulus period) in ∼70% of the C2Hx animals. We conclude that medullary inspiratory output can be used to trigger cervical ISMS and diaphragm activity after SCI. Further refinement of this method may enable "closed-loop-like" ISMS approaches to sustain ventilation after severe SCI.NEW & NOTEWORTHY We examined the feasibility of using intraspinal microstimulation (ISMS) of the cervical spinal cord to evoke diaphragm activity ipsilateral to acute and subacute hemisection of the upper cervical spinal cord of the rat. This proof-of-concept study demonstrated the efficacy of diaphragm activation, using an upper airway respiratory EMG signal to trigger ISMS at the level of the ipsilesional phrenic nucleus during acute and advanced postinjury intervals.

Coupling multielectrode array recordings with silver labeling of recording sites to study cervical spinal network connectivity.

Midcervical spinal interneurons form a complex and diffuse network and may be involved in modulating phrenic motor output. The intent of the current work was to enable a better understanding of midcervical "network-level" connectivity by pairing the neurophysiological multielectrode array (MEA) data with histological verification of the recording locations. We first developed a method to deliver 100-nA currents to electroplate silver onto and subsequently deposit silver from electrode tips after obtaining midcervical (C3-C5) recordings using an MEA in anesthetized and ventilated adult rats. Spinal tissue was then fixed, harvested, and histologically processed to "develop" the deposited silver. Histological studies verified that the silver deposition method discretely labeled (50-µm resolution) spinal recording locations between laminae IV and X in cervical segments C3-C5. Using correlative techniques, we next tested the hypothesis that midcervical neuronal discharge patterns are temporally linked. Cross-correlation histograms produced few positive peaks (5.3%) in the range of 0-0.4 ms, but 21.4% of neuronal pairs had correlogram peaks with a lag of ≥0.6 ms. These results are consistent with synchronous discharge involving mono- and polysynaptic connections among midcervical neurons. We conclude that there is a high degree of synaptic connectivity in the midcervical spinal cord and that the silver-labeling method can reliably mark metal electrode recording sites and "map" interneuron populations, thereby providing a low-cost and effective tool for use in MEA experiments. We suggest that this method will be useful for further exploration of midcervical network connectivity.NEW & NOTEWORTHY We describe a method that reliably identifies the locations of multielectrode array (MEA) recording sites while preserving the surrounding tissue for immunohistochemistry. To our knowledge, this is the first cost-effective method to identify the anatomic locations of neuronal ensembles recorded with a MEA during acute preparations without the requirement of specialized array electrodes. In addition, evaluation of activity recorded from silver-labeled sites revealed a previously unappreciated degree of connectivity between midcervical interneurons.

Transcriptome assessment of the Pompe (Gaa-/-) mouse spinal cord indicates widespread neuropathology.

Pompe disease, caused by deficiency of acid alpha-glucosidase (GAA), leads to widespread glycogen accumulation and profound neuromuscular impairments. There has been controversy, however, regarding the role of central nervous system pathology in Pompe motor dysfunction. We hypothesized that absence of GAA protein causes progressive activation of neuropathological signaling, including pathways associated with cell death. To test this hypothesis, genomic data (Affymetrix Mouse Gene Array 2.0ST) from the midcervical spinal cord in 6 and 16 mo old Pompe (Gaa-/-) mice were evaluated (Broad Institute Molecular Signature Database), along with spinal cord histology. The midcervical cord was selected because it contains phrenic motoneurons, and phrenic-diaphragm dysfunction is prominent in Pompe disease. Several clinically important themes for the neurologic etiology of Pompe disease emerged from this unbiased genomic assessment. First, pathways associated with cell death were strongly upregulated as Gaa-/- mice aged, and motoneuron apoptosis was histologically verified. Second, proinflammatory signaling was dramatically upregulated in the Gaa-/- spinal cord. Third, many signal transduction pathways in the Gaa-/- cervical cord were altered in a manner suggestive of impaired synaptic function. Notably, glutamatergic signaling pathways were downregulated, as were "synaptic plasticity pathways" including genes related to neuroplasticity. Fourth, many genes and pathways related to cellular metabolism are dysregulated. Collectively, the data unequivocally confirm that systemic absence of GAA induces a complex neuropathological cascade in the spinal cord. Most importantly, the results indicate that Pompe is a neurodegenerative condition, and this underscores the need for early therapeutic intervention capable of targeting the central nervous system.

Ampakine CX717 potentiates intermittent hypoxia-induced hypoglossal long-term facilitation.

Glutamatergic currents play a fundamental role in regulating respiratory motor output and are partially mediated by α-amino-3-hydroxy-5-methyl-isoxazole-propionic acid (AMPA) receptors throughout the premotor and motor respiratory circuitry. Ampakines are pharmacological compounds that enhance glutamatergic transmission by altering AMPA receptor channel kinetics. Here, we examined if ampakines alter the expression of respiratory long-term facilitation (LTF), a form of neuroplasticity manifested as a persistent increase in inspiratory activity following brief periods of reduced O2 [intermittent hypoxia (IH)]. Current synaptic models indicate enhanced effectiveness of glutamatergic synapses after IH, and we hypothesized that ampakine pretreatment would potentiate IH-induced LTF of respiratory activity. Inspiratory bursting was recorded from the hypoglossal nerve of anesthetized and mechanically ventilated mice. During baseline (BL) recording conditions, burst amplitude was stable for at least 90 min (98 ± 5% BL). Exposure to IH (3 × 1 min, 15% O2) resulted in a sustained increase in burst amplitude (218 ± 44% BL at 90 min following final bout of hypoxia). Mice given an intraperitoneal injection of ampakine CX717 (15 mg/kg) 10 min before IH showed enhanced LTF (500 ± 110% BL at 90 min). Post hoc analyses indicated that CX717 potentiated LTF only when initial baseline burst amplitude was low. We conclude that under appropriate conditions ampakine pretreatment can potentiate IH-induced respiratory LTF. These data suggest that ampakines may have therapeutic value in the context of hypoxia-based neurorehabilitation strategies, particularly in disorders with blunted respiratory motor output such as spinal cord injury.

Multicenter Evaluation of BD Max Enteric Parasite Real-Time PCR Assay for Detection of Giardia duodenalis, Cryptosporidium hominis, Cryptosporidium parvum, and Entamoeba histolytica.

Common causes of chronic diarrhea among travelers worldwide include protozoan parasites. The majority of parasitic infections are caused by Giardia duodenalis, Entamoeba histolytica, Cryptosporidium parvum, and Cryptosporidium hominis Similarly, these species cause the majority of parasitic diarrhea acquired in the United States. Detection of parasites by gold standard microscopic methods is time-consuming and requires considerable expertise; enzyme immunoassays and direct fluorescent-antibody (DFA) stains have lowered hands-on time for testing, but improvements in sensitivity and technical time may be possible with a PCR assay. We performed a clinical evaluation of a multiplex PCR panel, the enteric parasite panel (EPP), for the detection of these common parasites using the BD Max instrument, which performs automated extraction and amplification. A total of 2,495 compliant specimens were enrolled, including 2,104 (84%) specimens collected prospectively and 391 (16%) specimens collected retrospectively. Approximately equal numbers were received in 10% formalin (1,273 specimens) and unpreserved (1,222 specimens). The results from the EPP were compared to those from alternate PCR and bidirectional sequencing (APCR), as well as DFA (G. duodenalis and C. parvum or C. hominis) or trichrome stain (E. histolytica). The sensitivity and specificity for prospective and retrospective specimens combined were 98.2% and 99.5% for G. duodenalis, 95.5% and 99.6 for C. parvum or C. hominis, and 100% and 100% for E. histolytica, respectively. The performance of the FDA-approved BD Max EPP compared well to the reference methods and may be an appropriate substitute for microscopic examination or immunoassays.

Midcervical neuronal discharge patterns during and following hypoxia.

Anatomical evidence indicates that midcervical interneurons can be synaptically coupled with phrenic motoneurons. Accordingly, we hypothesized that interneurons in the C3-C4 spinal cord can display discharge patterns temporally linked with inspiratory phrenic motor output. Anesthetized adult rats were studied before, during, and after a 4-min bout of moderate hypoxia. Neuronal discharge in C3-C4 lamina I-IX was monitored using a multielectrode array while phrenic nerve activity was extracellularly recorded. For the majority of cells, spike-triggered averaging (STA) of ipsilateral inspiratory phrenic nerve activity based on neuronal discharge provided no evidence of discharge synchrony. However, a distinct STA phrenic peak with a 6.83 ± 1.1 ms lag was present for 5% of neurons, a result that indicates a monosynaptic connection with phrenic motoneurons. The majority (93%) of neurons changed discharge rate during hypoxia, and the diverse responses included both increased and decreased firing. Hypoxia did not change the incidence of STA peaks in the phrenic nerve signal. Following hypoxia, 40% of neurons continued to discharge at rates above prehypoxia values (i.e., short-term potentiation, STP), and cells with initially low discharge rates were more likely to show STP (P < 0.001). We conclude that a population of nonphrenic C3-C4 neurons in the rat spinal cord is synaptically coupled to the phrenic motoneuron pool, and these cells can modulate inspiratory phrenic output. In addition, the C3-C4 propriospinal network shows a robust and complex pattern of activation both during and following an acute bout of hypoxia.

Comparison of the Vidas C. difficile GDH Automated Enzyme-Linked Fluorescence Immunoassay (ELFA) with Another Commercial Enzyme Immunoassay (EIA) (Quik Chek-60), Two Selective Media, and a PCR Assay for gluD for Detection of Clostridium difficile in Fecal Samples.

Prevention and management of Clostridium difficile infection (CDI) can be improved by rapid and reliable diagnostics. The Vidas C. difficile glutamate dehydrogenase assay had performance comparable to that of the Quik Chek-60 assay (overall agreement, 95%) and a sensitivity of >93%; thus, it is suitable as the first test in two-stage algorithms for a CDI diagnosis.

Disruption of MBD5 contributes to a spectrum of psychopathology and neurodevelopmental abnormalities.

Microdeletions of chromosomal region 2q23.1 that disrupt MBD5 (methyl-CpG-binding domain protein 5) contribute to a spectrum of neurodevelopmental phenotypes; however, the impact of this locus on human psychopathology has not been fully explored. To characterize the structural variation landscape of MBD5 disruptions and the associated human psychopathology, 22 individuals with genomic disruption of MBD5 (translocation, point mutation and deletion) were identified through whole-genome sequencing or cytogenomic microarray at 11 molecular diagnostic centers. The genomic impact ranged from a single base pair to 5.4 Mb. Parents were available for 11 cases, all of which confirmed that the rearrangement arose de novo. Phenotypes were largely indistinguishable between patients with full-segment 2q23.1 deletions and those with intragenic MBD5 rearrangements, including alterations confined entirely to the 5'-untranslated region, confirming the critical impact of non-coding sequence at this locus. We identified heterogeneous, multisystem pathogenic effects of MBD5 disruption and characterized the associated spectrum of psychopathology, including the novel finding of anxiety and bipolar disorder in multiple patients. Importantly, one of the unique features of the oldest known patient was behavioral regression. Analyses also revealed phenotypes that distinguish MBD5 disruptions from seven well-established syndromes with significant diagnostic overlap. This study demonstrates that haploinsufficiency of MBD5 causes diverse phenotypes, yields insight into the spectrum of resulting neurodevelopmental and behavioral psychopathology and provides clinical context for interpretation of MBD5 structural variations. Empirical evidence also indicates that disruption of non-coding MBD5 regulatory regions is sufficient for clinical manifestation, highlighting the limitations of exon-focused assessments. These results suggest an ongoing perturbation of neurological function throughout the lifespan, including risks for neurobehavioral regression.