Spatial methane pattern in a deep freshwater lake: Relation to water depth and topography.

Freshwater lakes are regarded as important methane (CH4) sources, accounting for ~20% of natural emission. To improve the assessment of the global greenhouse effect, it is necessary to consider spatial variability within lakes. Here, CH4 concentrations in the water column and sediment layers, as well as the sediment CH4 production potentials and diffusive fluxes, were studied in the littoral, intermediate, and profundal zones of the medium-sized (425 ha), deep (maximum depth 69.5 m) Lake Stechlin (Germany). Sediment CH4 concentrations, production potentials and sediment-water interface diffusive fluxes showed significant spatial heterogeneity and were highest in the profundal zone. CH4 concentrations in the surface water did not differ among the studied locations, indicating a decoupling from the production sites in the sediment. The high amount of CH4 in profundal sediments that might potentially be released to the atmosphere is either trapped or oxidized within the water column, while the surface water dissolved CH4 is more related to the dynamics in the epilimnion. The divergence in sediment physical (water content, grain size) and chemical (organic matter quantity or quality, sulfate) properties across the lake leads to variations in CH4 dynamics which are restricted to deeper habitats in this type of lake.

Spread of Terbinafine-Resistant Trichophyton mentagrophytes Type VIII (India) in Germany-"The Tip of the Iceberg?"

Chronic recalcitrant dermatophytoses, due to Trichophyton (T.) mentagrophytes Type VIII are on the rise in India and are noteworthy for their predominance. It would not be wrong to assume that travel and migration would be responsible for the spread of T. mentagrophytes Type VIII from India, with many strains resistant to terbinafine, to other parts of the world. From September 2016 until March 2020, a total of 29 strains of T. mentagrophytes Type VIII (India) were isolated. All patients were residents of Germany: 12 females, 15 males and the gender of the remaining two was not assignable. Patients originated from India (11), Pakistan (two), Bangladesh (one), Iraq (two), Bahrain (one), Libya (one) and other unspecified countries (10). At least two patients were German-born residents. Most samples (21) were collected in 2019 and 2020. All 29 T. mentagrophytes isolates were sequenced (internal transcribed spacer (ITS) and translation elongation factor 1-α gene (TEF1-α)). All were identified as genotype VIII (India) of T. mentagrophytes. In vitro resistance testing revealed 13/29 strains (45%) to be terbinafine-resistant with minimum inhibitory concentration (MIC) breakpoints ≥0.2 µg/mL. The remaining 16 strains (55%) were terbinafine-sensitive. Point mutation analysis revealed that 10/13 resistant strains exhibited Phe397Leu amino acid substitution of squalene epoxidase (SQLE), indicative for in vitro resistance to terbinafine. Two resistant strains showed combined Phe397Leu and Ala448Thr amino acid substitutions, and one strain a single Leu393Phe amino acid substitution. Out of 16 terbinafine-sensitive strains, in eight Ala448Thr, and in one Ala448Thr +, new Val444 Ile amino acid substitutions were detected. Resistance to both itraconazole and voriconazole was observed in three out of 13 analyzed strains. Treatment included topical ciclopirox olamine plus topical miconazole or sertaconazole. Oral itraconazole 200 mg twice daily for four to eight weeks was found to be adequate. Terbinafine-resistant T. mentagrophytes Type VIII are being increasingly isolated. In Germany, transmission of T. mentagrophytes Type VIII from the Indian subcontinent to Europe should be viewed as a significant public health issue.

Early delivery and prolonged treatment with nimodipine prevents the development of spasticity after spinal cord injury in mice.

Spasticity, one of the most frequent comorbidities of spinal cord injury (SCI), disrupts motor recovery and quality of life. Despite major progress in neurorehabilitative and pharmacological approaches, therapeutic strategies for treating spasticity are lacking. Here, we show in a mouse model of chronic SCI that treatment with nimodipine-an L-type calcium channel blocker already approved from the European Medicine Agency and from the U.S. Food and Drug Administration-starting in the acute phase of SCI completely prevents the development of spasticity measured as increased muscle tone and spontaneous spasms. The aberrant muscle activities associated with spasticity remain inhibited even after termination of the treatment. Constitutive and conditional silencing of the L-type calcium channel CaV1.3 in neuronal subtypes demonstrated that this channel mediated the preventive effect of nimodipine on spasticity after SCI. This study identifies a treatment protocol and suggests that targeting CaV1.3 could prevent spasticity after SCI.

Modeling Motor Neuron Resilience in ALS Using Stem Cells.

Oculomotor neurons, which regulate eye movement, are resilient to degeneration in the lethal motor neuron disease amyotrophic lateral sclerosis (ALS). It would be highly advantageous if motor neuron resilience could be modeled in vitro. Toward this goal, we generated a high proportion of oculomotor neurons from mouse embryonic stem cells through temporal overexpression of PHOX2A in neuronal progenitors. We demonstrate, using electrophysiology, immunocytochemistry, and RNA sequencing, that in vitro-generated neurons are bona fide oculomotor neurons based on their cellular properties and similarity to their in vivo counterpart in rodent and man. We also show that in vitro-generated oculomotor neurons display a robust activation of survival-promoting Akt signaling and are more resilient to the ALS-like toxicity of kainic acid than spinal motor neurons. Thus, we can generate bona fide oculomotor neurons in vitro that display a resilience similar to that seen in vivo.

Shifts among Eukaryota, Bacteria, and Archaea define the vertical organization of a lake sediment.

BACKGROUND: Lake sediments harbor diverse microbial communities that cycle carbon and nutrients while being constantly colonized and potentially buried by organic matter sinking from the water column. The interaction of activity and burial remained largely unexplored in aquatic sediments. We aimed to relate taxonomic composition to sediment biogeochemical parameters, test whether community turnover with depth resulted from taxonomic replacement or from richness effects, and to provide a basic model for the vertical community structure in sediments. METHODS: We analyzed four replicate sediment cores taken from 30-m depth in oligo-mesotrophic Lake Stechlin in northern Germany. Each 30-cm core spanned ca. 170 years of sediment accumulation according to 137Cs dating and was sectioned into layers 1-4 cm thick. We examined a full suite of biogeochemical parameters and used DNA metabarcoding to examine community composition of microbial Archaea, Bacteria, and Eukaryota. RESULTS: Community ß-diversity indicated nearly complete turnover within the uppermost 30 cm. We observed a pronounced shift from Eukaryota- and Bacteria-dominated upper layers (<5 cm) to Bacteria-dominated intermediate layers (5-14 cm) and to deep layers (>14 cm) dominated by enigmatic Archaea that typically occur in deep-sea sediments. Taxonomic replacement was the prevalent mechanism in structuring the community composition and was linked to parameters indicative of microbial activity (e.g., CO2 and CH4 concentration, bacterial protein production). Richness loss played a lesser role but was linked to conservative parameters (e.g., C, N, P) indicative of past conditions. CONCLUSIONS: By including all three domains, we were able to directly link the exponential decay of eukaryotes with the active sediment microbial community. The dominance of Archaea in deeper layers confirms earlier findings from marine systems and establishes freshwater sediments as a potential low-energy environment, similar to deep sea sediments. We propose a general model of sediment structure and function based on microbial characteristics and burial processes. An upper "replacement horizon" is dominated by rapid taxonomic turnover with depth, high microbial activity, and biotic interactions. A lower "depauperate horizon" is characterized by low taxonomic richness, more stable "low-energy" conditions, and a dominance of enigmatic Archaea.

Spatiotemporal correlation of spinal network dynamics underlying spasms in chronic spinalized mice.

Spasms after spinal cord injury (SCI) are debilitating involuntary muscle contractions that have been associated with increased motor neuron excitability and decreased inhibition. However, whether spasms involve activation of premotor spinal excitatory neuronal circuits is unknown. Here we use mouse genetics, electrophysiology, imaging and optogenetics to directly target major classes of spinal interneurons as well as motor neurons during spasms in a mouse model of chronic SCI. We find that assemblies of excitatory spinal interneurons are recruited by sensory input into functional circuits to generate persistent neural activity, which interacts with both the graded expression of plateau potentials in motor neurons to generate spasms, and inhibitory interneurons to curtail them. Our study reveals hitherto unrecognized neuronal mechanisms for the generation of persistent neural activity under pathophysiological conditions, opening up new targets for treatment of muscle spasms after SCI.

Descending Command Neurons in the Brainstem that Halt Locomotion.

The episodic nature of locomotion is thought to be controlled by descending inputs from the brainstem. Most studies have largely attributed this control to initiating excitatory signals, but little is known about putative commands that may specifically determine locomotor offset. To link identifiable brainstem populations to a potential locomotor stop signal, we used developmental genetics and considered a discrete neuronal population in the reticular formation: the V2a neurons. We find that those neurons constitute a major excitatory pathway to locomotor areas of the ventral spinal cord. Selective activation of V2a neurons of the rostral medulla stops ongoing locomotor activity, owing to an inhibition of premotor locomotor networks in the spinal cord. Moreover, inactivation of such neurons decreases spontaneous stopping in vivo. Therefore, the V2a "stop neurons" represent a glutamatergic descending pathway that favors immobility and may thus help control the episodic nature of locomotion.

Selective mitochondrial Ca2+ uptake deficit in disease endstage vulnerable motoneurons of the SOD1G93A mouse model of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis is a progressive neurodegenerative disease that targets some somatic motoneuron populations, while others, e.g. those of the oculomotor system, are spared. The pathophysiological basis of this pattern of differential vulnerability, which is preserved in a transgenic mouse model of amyotrophic lateral sclerosis (SOD1(G93A)), and the mechanism of neurodegeneration in general are unknown. Hyperexcitability and calcium dysregulation have been proposed by others on the basis of data from juvenile mice that are, however, asymptomatic. No studies have been done with symptomatic mice following disease progression to the disease endstage. Here, we developed a new brainstem slice preparation for whole-cell patch-clamp recordings and single cell fura-2 calcium imaging to study motoneurons in adult wild-type and SOD1(G93A) mice up to disease endstage. We analysed disease-stage-dependent electrophysiological properties and intracellular Ca(2+) handling of vulnerable hypoglossal motoneurons in comparison to resistant oculomotor neurons. Thereby, we identified a transient hyperexcitability in presymptomatic but not in endstage vulnerable motoneurons. Additionally, we revealed a remodelling of intracellular Ca(2+) clearance within vulnerable but not resistant motoneurons at disease endstage characterised by a reduction of uniporter-dependent mitochondrial Ca(2+) uptake and enhanced Ca(2+) extrusion across the plasma membrane. Our study challenged the notion that hyperexcitability is a direct cause of neurodegeneration in SOD1(G93A) mice, but molecularly identified a Ca(2+) clearance deficit in motoneurons and an adaptive Ca(2+) handling strategy that might be targeted by future therapeutic strategies.

Downregulation of the potassium chloride cotransporter KCC2 in vulnerable motoneurons in the SOD1-G93A mouse model of amyotrophic lateral sclerosis.

The balance between excitatory and inhibitory synaptic inputs is critical for the physiological control of motoneurons. The maintenance of a low-intracellular chloride concentration by the potassium chloride cotransporter 2 (KCC2) is essential for the efficacy of fast synaptic inhibition of mature motoneurons in response to the activation of ionotropic γ-aminobutyric acid A and glycine receptors. Altered synaptic balance and excitotoxicity have been proposed as candidate pathophysiological processes in amyotrophic lateral sclerosis (ALS). Therefore, we investigated the expression patterns of KCC2 and its functional opponent, the chloride influx-mediating sodium-potassium chloride cotransporter 1 (NKCC1), in the superoxide dismutase 1 (SOD1-G93A) mouse model of ALS. We detected reduced KCC2 messenger RNA levels and less membrane-bound KCC2 immunoreactivity in ALS-vulnerable motoneurons in lumbar spinal cord and hypoglossal nuclei of SOD1-G93A mice but not in degeneration-resistant oculomotor nuclei. Downregulation of KCC2 started during late presymptomatic stages and accelerated in parallel to hind limb and tongue motor function deficits. In contrast, NKCC1 messenger RNA levels were unaltered in postnatal lumbar spinal cord motoneurons. Our data indicate that reductions in KCC2 gene expression may contribute to selective motor deficits and disease progression in vulnerable motoneurons in a mouse model of ALS.

Validation of endogenous controls for quantitative gene expression analysis: application on brain cortices of human chronic alcoholics.

Real-time PCR is frequently used for gene expression quantification due to its methodological sensitivity and reproducibility. The gene expression is quantified by normalization to one or more reference genes, usually beta-actin (ACTB), glyceraldehyde-3-phosphate dehydrogenase (GAPD) or to ribosomal RNA (18S). However, different environmental or pathological conditions might also influence the expression of normalizing genes, which could severely skew the interpretation of quantitative results. This study evaluates whether 16 genes frequently used as endogenous controls in expression studies, can serve as such for comparison of human brain tissues of chronic alcoholics and control subjects. The prefrontal and motor cortices that are affected differently by chronic alcohol consumption were analyzed. The reference genes that have no or small differences in expression in alcoholics and control subjects, were found to be specific for each region: beta-actin (ACTB) and ribosomal large P0 (RPLP0) for the prefrontal cortex while importin 8 (IPO8) and RNA polymerase II (POLR2A) for the motor cortex. Four out of sixteen analyzed genes demonstrated significant differences in expression between alcoholics and controls: phosphoglycerate kinase (PGK1), hypoxanthine phosphoribosyl transferase (HPRT1) and peptidylprolyl isomerase A (PPIA) in the motor cortex and beta-2-microglobulin (B2M) in the prefrontal cortex. Our study demonstrates the importance of validation of endogenous control genes prior to real-time PCR analysis of human brain tissues. Prescribed and non-prescribed drugs, pathological or environmental conditions along with alcohol abuse may differentially influence expression of reference genes.