Background. Tenofovir disoproxil fumarate (TDF) is an established nucleotide analogue in the treatment of chronic hepatitis B. Bone mineral density loss has been described in TDF-treated patients ...with human immunodeficiency virus infection, but limited data exist for patients with chronic hepatitis B. Dual X-ray absorptiometry (DEXA) was used to determine bone mineral density changes in TDF-exposed patients. We evaluated the accuracy of the Fracture Risk Assessment Tool (FRAX) as an alternative to DEXA in clinical practice. Methods. A total of 170 patients were studied: 122 were exposed to TDF, and 48 were controls. All patients underwent DEXA, and demographic details were recorded. FRAX scores (before and after DEXA) were calculated. Results. TDF was associated with a lower hip T score (P = .02). On univariate and multivariate analysis, advancing age, smoking, lower body mass index, and TDF exposure were independent predictors of low bone mineral density. In addition, the pre-DEXA FRAX score was an accurate predictor of the post-DEXA FRAX treatment recommendation (100% sensitivity and 83% specificity), area under the curve 0.93 (95% CI, .87-.97, P<.001). Conclusions. TDF-treated patients with chronic hepatitis have reduced bone mineral density, but the reduction is limited to 1 anatomical site. Age and advanced liver disease are additional contributing factors, underlining the importance of multifactorial fracture risk assessment. FRAX can accurately identify those at greatest risk of osteoporotic fracture.
The mechanisms that facilitate animal magnetoreception have both fascinated and confounded scientists for decades, and its precise biophysical origin remains unclear. Among the proposed primary ...magnetic sensors is the flavoprotein, cryptochrome, which is thought to provide geomagnetic information via a quantum effect in a light-initiated radical pair reaction. Despite recent advances in the radical pair model of magnetoreception from theoretical, molecular and animal behaviour studies, very little is known of a possible signal transduction mechanism. We report a substantial effect of magnetic field exposure on seizure response in Drosophila larvae. The effect is dependent on cryptochrome, the presence and wavelength of light and is blocked by prior ingestion of typical antiepileptic drugs. These data are consistent with a magnetically-sensitive, photochemical radical pair reaction in cryptochrome that alters levels of neuronal excitation, and represent a vital step forward in our understanding of the signal transduction mechanism involved in animal magnetoreception.
Activity of voltage-gated Na channels (Na(v)) is modified by alternative splicing. However, whether altered splicing of human Na(v)s contributes to epilepsy remains to be conclusively shown. We show ...here that altered splicing of the Drosophila Na(v) (paralytic, DmNa(v)) contributes to seizure-like behavior in identified seizure mutants. We focus attention on a pair of mutually exclusive alternate exons (termed K and L), which form part of the voltage sensor (S4) in domain III of the expressed channel. The presence of exon L results in a large, non-inactivating, persistent I(Nap). Many forms of human epilepsy are associated with an increase in this current. In wild-type (WT) Drosophila larvae, ∼70-80% of DmNa(v) transcripts contain exon L, and the remainder contain exon K. Splicing of DmNa(v) to include exon L is increased to ∼100% in both the slamdance and easily-shocked seizure mutants. This change to splicing is prevented by reducing synaptic activity levels through exposure to the antiepileptic phenytoin or the inhibitory transmitter GABA. Conversely, enhancing synaptic activity in WT, by feeding of picrotoxin is sufficient to increase I(Nap) and promote seizure through increased inclusion of exon L to 100%. We also show that the underlying activity-dependent mechanism requires the presence of Pasilla, an RNA-binding protein. Finally, we use computational modeling to show that increasing I(Nap) is sufficient to potentiate membrane excitability consistent with a seizure phenotype. Thus, increased synaptic excitation favors inclusion of exon L, which, in turn, further increases neuronal excitability. Thus, at least in Drosophila, this self-reinforcing cycle may promote the incidence of seizure.
Studying ion channel currents generated distally from the recording site is difficult because of artifacts caused by poor space clamp and membrane filtering. A computational model can quantify ...artifact parameters for correction by simulating the currents only if their exact anatomical location is known. We propose that the same artifacts that confound current recordings can help pinpoint the source of those currents by providing a signature of the neuron's morphology. This method can improve the recording quality of currents initiated at the spike initiation zone (SIZ) that are often distal to the soma in invertebrate neurons. Drosophila being a valuable tool for characterizing ion currents, we estimated the SIZ location and quantified artifacts in an identified motoneuron, aCC/MN1-Ib, by constructing a novel multicompartmental model. Initial simulation of the measured biophysical channel properties in an isopotential Hodgkin-Huxley type neuron model partially replicated firing characteristics. Adding a second distal compartment, which contained spike-generating Na+ and K+ currents, was sufficient to simulate aCC's in vivo activity signature. Matching this signature using a reconstructed morphology predicted that the SIZ is on aCC's primary axon, 70 μm after the most distal dendritic branching point. From SIZ to soma, we observed and quantified selective morphological filtering of fast activating currents. Non-inactivating K+ currents are filtered ∼3 times less and despite their large magnitude at the soma they could be as distal as Na+ currents. The peak of transient component (NaT) of the voltage-activated Na+ current is also filtered more than the magnitude of slower persistent component (NaP), which can contribute to seizures. The corrected NaP/NaT ratio explains the previously observed discrepancy when the same channel is expressed in different cells. In summary, we used an in vivo signature to estimate ion channel location and recording artifacts, which can be applied to other neurons.
Epstein‐Barr virus (EBV) is part of the herpesvirus family that infects up to 90% of the population. Initial infection is often subclincal in children but will generally result in symptomatic ...infectious mononucleosis in adolescents and adults. Ganciclovir has been utilized in immunocompromised patients with EBV encephalitis and post‐liver transplant for EBV fulminant hepatitis. Herein, the successful use of ganciclovir in two immunocompetent patients with severe EBV hepatitis is reported.
The fruit fly Drosophila melanogaster has been instrumental in expanding our understanding of early aspects of neural development. The use of this model system has greatly added to our knowledge of ...neural cell-fate determination, axon guidance, and synapse formation. It has also become possible to access and make electrophysiological recordings directly from neurons in situ in an intact central nervous system (CNS), which has facilitated studies of the development and regulation of neuronal signaling. It is possible to obtain electrophysiological recordings from all stages of Drosophila. Exposure of the intact Drosophila CNS is a prerequisite for such electrophysiological recordings. The dissection procedure described here is suitable for third-instar larvae. The dissection should take ∼5 min to complete if all preparation work has been completed in advance. Owing to the short life span of the dissected larva, it is not recommended that the procedure be stopped or the preparation stored for later use.
There is clinical need to extend the understanding of epilepsy and to find novel approaches to treat this condition. Bang-sensitive (bs) Drosophila mutants, which exhibit reduced thresholds for ...seizure, offer an attractive possibility to combine tractable genetics, electrophysiology, and high-throughput screening. However, despite these advantages, the precise electrophysiological aberrations that contribute to seizure have not been identified in any bs mutant. Because of this, the applicability of Drosophila as a preclinical model has not yet been established. In this study, we show that electroshock of bs slamdance (sda) larvae was sufficient to induce extended seizure-like episodes. Whole cell voltage-clamp recordings from identified motoneurons (termed aCC and RP2) showed synaptic currents that were greatly increased in both amplitude and duration. Current-clamp recordings indicated that these inputs produced longer-lived plateau depolarizations and increased action potential firing in these cells. An analysis of voltage-gated currents in these motoneurons, in both first and third instar larvae, revealed a consistently increased persistent Na(+) current (I(Nap)) and a reduced Ca(2+) current in first instar larvae, which appeared normal in older third instar larvae. That increased I(Nap) may contribute to seizure-like activity is indicated by the observation that feeding sda larvae the antiepileptic drug phenytoin, which was sufficient to reduce I(Nap), rescued both seizure-like episode duration and synaptic excitation of motoneurons. In contrast, feeding of either anemone toxin, a drug that preferentially increases I(Nap), or phenytoin to wild-type larvae was sufficient to induce a bs behavioral phenotype. Finally, we show that feeding of phenytoin to gravid sda females was sufficient to both reduce I(Nap) and synaptic currents and rescue the bs phenotype in their larval progeny, indicating that a heightened predisposition to seizure may arise as a consequence of abnormal embryonic neural development.
Multiple neuropeptides are known to regulate water and ion balance in
Drosophila melanogaster
. Several of these peptides also have other functions in physiology and behavior. Examples are ...corticotropin-releasing factor-like diuretic hormone (diuretic hormone 44; DH44) and leucokinin (LK), both of which induce fluid secretion by Malpighian tubules (MTs), but also regulate stress responses, feeding, circadian activity and other behaviors. Here, we investigated the functional relations between the LK and DH44 signaling systems. DH44 and LK peptides are only colocalized in a set of abdominal neurosecretory cells (ABLKs). Targeted knockdown of each of these peptides in ABLKs leads to increased resistance to desiccation, starvation and ionic stress. Food ingestion is diminished by knockdown of DH44, but not LK, and water retention is increased by LK knockdown only. Thus, the two colocalized peptides display similar systemic actions, but differ with respect to regulation of feeding and body water retention. We also demonstrated that DH44 and LK have additive effects on fluid secretion by MTs. It is likely that the colocalized peptides are coreleased from ABLKs into the circulation and act on the tubules where they target different cell types and signaling systems to regulate diuresis and stress tolerance. Additional targets seem to be specific for each of the two peptides and subserve regulation of feeding and water retention. Our data suggest that the ABLKs and hormonal actions are sufficient for many of the known DH44 and LK functions, and that the remaining neurons in the CNS play other functional roles.
Abstract A neuromodulatory role for nitric oxide has been reported for magnocellular neuroendocrine cells in mammalian hypothalamus. We examined its potential as a local intercellular messenger in ...the neuroendocrine Dahlgren cell population of the caudal neurosecretory system (CNSS) of the euryhaline flounder. Immunocytochemistry using an antibody raised against human neuronal nitric oxide synthase (NOS) indicated the presence of NOS in the Dahlgren cells. Quantitative RT-PCR, using a flounder-specific probe, revealed NOS mRNA expression in the CNSS. In July, though not in September, NOS mRNA expression was significantly higher in fish fully adapted to seawater, compared to freshwater-adapted fish. Following acute transfer of fish from freshwater to seawater, NOS mRNA expression was elevated at 8 h and then recovered by 24 h. In pharmacological experiments in vitro , application of NO donors (SNAP, SNP) caused an increase in electrical activity (firing frequency) of Dahlgren cells, recruitment of previously silent cells, together with a greater proportion of cells showing phasic (irregular) activity. The NOS substrate, l -arginine, led to increased firing frequency, cell recruitment and enhanced bursting activity. However, this effect was not blocked by the NOS inhibitor L-NAME. These findings suggest that NO acts as a modulator within the CNSS, potentially enhancing electrical activity and hence secretory output. A role in supporting adaptation to hyperosmotic conditions is also indicated.
Sepsis is a common complication of cirrhosis with a high mortality. In this study, we have investigated some of the pathways that may be involved in tissue injury and death. Bile duct–ligated (BDL) ...cirrhotic and control rats were challenged with lipopolysaccharide (LPS). Sensitivity to LPS was markedly enhanced in the BDL group, and was associated with increased liver injury and mortality. There was a 5‐fold constitutive activation of nuclear factor κ B (NFκB) in the liver of BDL rat controls (P < .001), and this was activated further, but to a similar extent, in the liver of both sham and BDL rats after injection of LPS. Plasma tumor necrosis factor α (TNF‐α) increased more markedly in the BDL cirrhotic rats (2,463 ± 697 pg/mL in BDL rats versus 401 ± 160 pg/mL in the controls at 3 hours;
P < .01). Plasma nitrite/nitrate concentrations were increased in the BDL controls at baseline, and increased further after LPS (P < .05), but did not differ from sham controls at 6 hours. Plasma F2
‐isoprostanes increased 6‐fold in the cirrhotic rats and 2‐fold in the controls (P < .01) indicative of lipid peroxidation. Esterified F2
‐isoprostanes in the liver increased 2‐ to 3‐fold at 1 hour in control and BDL rats, but returned to baseline levels by 3 hours. Esterified F2
‐isoprostanes in the kidney increased by 2‐fold in the BDL rats after LPS administration, but remained unchanged in sham controls. We conclude that there is a marked increase in sensitivity to LPS in BDL cirrhotic rats. This is associated with an enhanced TNF‐α response and increased lipid peroxidation. These may be directly and causally related to mortality.
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