One-stage surgical case management of a two-year-old Arabian horse affected by male-pseudo hermaphroditism.

A two-year-old Arabian horse presented for abnormal external genitalia and dangerous stallion-like behavior was diagnosed with disorder of sexual development (DSD), also known as intersex/hermaphroditism. Standing 1-stage surgical procedure performed under sedation, and local anesthesia to concurrently eliminate stallion-like behavior, risk of neoplastic transformation of intraabdominal gonads, and to replace ambiguous external genital with a functional, and cosmetically more acceptable anatomy. Step-1) Laparoscopic abdominal exploration and gonadectomy; Step-2) Rudimentary penis resection and perineal urethrostomy. The horse tolerated surgery well (combined surgery time 185 min) with no complications. At macroscopic examination of the gonads, they resembled hypoplastic testis-like tissues. Microscopic examination confirmed presence of seminiferous tubules, Leydig and Sertoli/granulosa cells. Cytogenetic evaluation revealed a 64,XX karyotype, SRY-negative. The stallion-like behavior subsided within days post-operatively. Long-term follow-up revealed the genitoplasty site healed without urine scalding or urethral stricture. The owner satisfaction was excellent and the horse could be used post-surgery as an athlete.

Structure-function relationship of new peptides activating human Nav1.1.

Nav1.1 is an important pharmacological target as this voltage-gated sodium channel is involved in neurological and cardiac syndromes. Channel activators are actively sought to try to compensate for haploinsufficiency in several of these pathologies. Herein we used a natural source of new peptide compounds active on ion channels and screened for drugs capable to inhibit channel inactivation as a way to compensate for decreased channel function. We discovered that JzTx-34 is highly active on Nav1.1 and subsequently performed a full structure-activity relationship investigation to identify its pharmacophore. These experiments will help interpret the mechanism of action of this and formerly identified peptides as well as the future identification of new peptides. We also reveal structural determinants that make natural ICK peptides active against Nav1.1 challenging to synthesize. Altogether, the knowledge gained by this study will help facilitate the discovery and development of new compounds active on this critical ion channel target.

Identification of CNS-Penetrant Aryl Sulfonamides as Isoform-Selective NaV1.6 Inhibitors with Efficacy in Mouse Models of Epilepsy.

Nonselective antagonists of voltage-gated sodium (NaV) channels have been long used for the treatment of epilepsies. The efficacy of these drugs is thought to be due to the block of sodium channels on excitatory neurons, primarily NaV1.6 and NaV1.2. However, these currently marketed drugs require high drug exposure and suffer from narrow therapeutic indices. Selective inhibition of NaV1.6, while sparing NaV1.1, is anticipated to provide a more effective and better tolerated treatment for epilepsies. In addition, block of NaV1.2 may complement the anticonvulsant activity of NaV1.6 inhibition. We discovered a novel series of aryl sulfonamides as CNS-penetrant, isoform-selective NaV1.6 inhibitors, which also displayed potent block of NaV1.2. Optimization focused on increasing selectivity over NaV1.1, improving metabolic stability, reducing active efflux, and addressing a pregnane X-receptor liability. We obtained compounds 30-32, which produced potent anticonvulsant activity in mouse seizure models, including a direct current maximal electroshock seizure assay.

Surgical management of primary spinal hemangiopericytomas: an institutional case series and review of the literature.

PURPOSE: Hemangiopericytoma (HPC) is a rare tumor of the central nervous system. Primary spinal occurrence of this tumor is extremely uncommon and cases involving the intramedullary spinal cord are even more rare. The purpose of this study was to explore the clinical features, surgical strategies, outcome and pathology in a consecutive series of patients treated at a single institution. METHODS: The authors performed a retrospective review of the clinicopathological characteristics of four patients with a pathological diagnosis of spinal HPC. RESULTS: Four cases with intradural as well as intra/extra-medullary components were identified. Gross total resection with no recurrence at the operative site was achieved in the majority of patients with a spinal HPC. One patient had significant recurrence and eventually, succumbed to the disease. CONCLUSION: Increased awareness of these tumors' capability to occur intradurally and intramedullarly can help surgeons accurately diagnose and choose an effective plan of care. Gross total resection of hemangiopericytomas is the mainstay of treatment and should be pursued if feasible. Histopathology is essential to the diagnosis.

Effect of fuel and lube oil sulfur on the performance of a diesel exhaust gas continuously regenerating trap.

The continuously regenerating trap (CRT) is a diesel exhaust emission control that removes nearly all diesel particulate matter on a mass basis, but under some circumstances oxidation of sulfur leads to the formation of nanoparticles. The objective of the four year study was to determine CRT performance under controlled, real-world, on-road conditions, and to develop quantitative relationships between fuel and lubrication oil sulfur concentration and particle number exhaust emissions. It was shown that nanoparticle emissions are minimized by the use of ultralow sulfur fuels and specially formulated low sulfur lubrication oil. Nanoparticle emissions increased with higher exhaust temperatures. Fuel and lubrication oil sulfur increased the particle concentration by, on average, 36 x 10(6) and 0.14 x 10(6) part/cm3 for each 1 ppm increase in sulfur. On the other hand there was a decrease in nanoparticle emissions by the CRT as the system aged.

The carboxyl-terminal region of cyclic nucleotide-modulated channels is a gating ring, not a permeation path.

The recent elucidation of the structure of the carboxyl-terminal region of the hyperpolarization-activated cyclic nucleotide-modulated (HCN2) channel has prompted us to investigate a curious feature of this structure in HCN2 channels and in the related CNGA1 cyclic nucleotide-gated (CNG) channels. The crystallized fragment of the HCN2 channel contains both the cyclic nucleotide-binding domain (CNBD) and the C-linker region, which connects the CNBD to the pore. At the center of the fourfold-symmetric structure is a tunnel that runs perpendicular to the membrane. The narrowest part of the tunnel is approximately 10 A in diameter and is lined by a ring of negatively charged amino acids: D487, E488, and D489. Many ion channels have "charge rings" that focus permeant ions at the mouth of the pore and increase channel conductance. We used nonstationary fluctuation analysis and single-channel recording, coupled with site-directed mutagenesis and cysteine modification, to determine whether this part of HCN and CNG channels might be an extension of the permeation pathway. Our results indicate that modifying charge-ring amino acids affects gating but not ion permeation in HCN2 and CNG channels. Thus, this portion of the channel is not an obligatory part of the ion path but instead acts as a "gating ring." The carboxyl-terminal region of these channels must hang below the pore much like the "hanging gondola" of voltage-gated potassium channels, but the permeation pathway must exit the protein before the level of the ring of charged amino acids.

On-road exposure to highway aerosols. 1. Aerosol and gas measurements.

On-road experiments were conducted to determine the sensitivities of rats to real-world aerosol. This article summarizes the on-road aerosol and gas measurements and provides background information for the companion paper on the rat exposures. Measurements were carried out over 10 days, 6 h/day, driving a route from Rochester to Buffalo. Aerosol instrumentation used in this study included two scanning mobility particle sizers (SMPS) to determine the aerosol size distribution from 10 to 300 nm, 2 stand-alone condensation particle counters to determine the total aerosol number concentration, and an electrical aerosol detector to determine the aerosol length concentration. A thermal denuder (TD) was used with one of the SMPS instruments to determine the size distribution of the non-volatile fraction. Filter samples were collected and analyzed for elemental carbon, and gas analyzers measured ambient levels of CO, CO(2), and NO. Average daily total aerosol number concentration ranged from 200,000 to 560,000 particles/cm(3). Past studies on urban highways have measured total number concentrations ranging between 10(4) and 10(6) particles/cm(3). The average daily NO concentration ranged from 0.10 to 0.24 ppm and the corresponding CO(2) concentration ranged from 400 to 420 ppm. The average daily geometric number mean particle size determined by the SMPS ranged from 15 to 20 nm. The TD reduced the average SMPS number concentration between 87 and 95% and the SMPS volume between 54 and 83%, suggesting that most of the particles consisted of volatile material. The TD also increased the geometric number mean diameter from 15 to 20 nm to 30 to 40 nm.

Frameless stereotaxy for anterior spinal procedures.

OBJECT: Intraoperative image guidance provides real-time three-dimensional visualization and has been successfully applied in many posterior spinal procedures. The feasibility of applying these techniques to anterior spinal surgery has not been studied systematically because the anterior spine, in contrast to the posterior spine, lacks distinct anatomical landmarks for registration. The authors sought to evaluate the practicality of performing stereotaxy in the anterior spine in a cadaveric model. METHODS: Unilateral C4-L4 pedicle screws were placed posteriorly in three cadaveric specimens to serve as unknown markers within each vertebral body. The specimens then underwent computerized tomography (CT) scanning, and the CT data were transferred to an optical tracking system. The anterior surface of the spine was registered for use with the stereotactic system by using a paired point-matching technique. Attached to a surgical drill, K-wires were placed under stereotactic guidance in a tip-to-tip orientation with the posterior pedicle screws. A second postoperative CT scan was obtained, and accuracy was determined by measuring the distance between the tips of the K-wire and pedicle screw. The K-wires were placed tip to tip with pedicle screw markers in 57 vertebral levels. The mean registration error was 1.47+/-0.04 mm, and when combined with the universal instrument registration error of 0.7 mm yielded an overall registration error of 2.17+/-0.04 mm. The mean tip-to-tip distance for all K-wires placed was 2.46+/-0.23 mm. The difference between the mean tip-to-tip distance and overall registration error was not statistically significant (p > 0.05), indicating that the K-wires were placed within the expected range of error. CONCLUSIONS: The results of this study confirmed the feasibility of performing anterior stereotactic procedures throughout the spine. The accuracy of the findings in this study indicates that anterior stereotaxy should be applicable in clinical practice.

A novel extracellular calcium sensing mechanism in voltage-gated potassium ion channels.

Potassium (K(+)) channels influence neurotransmitter release, burst firing rate activity, pacing, and critical dampening of neuronal circuits. Internal and external factors that further modify K(+) channel function permit fine-tuning of neuronal circuits. Human ether-à-go-go-related gene (HERG) K(+) channels are unusually sensitive to external calcium concentration ([Ca(2+)](o)). Small changes in [Ca(2+)](o) shift the voltage dependence of channel activation to more positive membrane potentials, an effect that cannot be explained by nonspecific surface charge screening or channel pore block. The HERG-calcium concentration-response relationship spans the physiological range for [Ca(2+)](o). The modulatory actions of calcium are attributable to differences in the Ca(2+) affinity between rested and activated channels. Adjacent extracellular, negatively charged amino acids (E518 and E519) near the S4 voltage sensor influence both channel gating and Ca(2+) dependence. Neutralization of these charges had distinct effects on channel gating and calcium sensitivity. A change in the degree of energetic coupling between these amino acids on transition from closed to activated channel states reveals movement in this region during channel gating and defines a molecular mechanism for protein state-dependent ligand interactions. The results suggest a novel extracellular [Ca(2+)](o) sensing mechanism coupled to allosteric changes in channel gating and a mechanism for fine-tuning cell repolarization.

Image-guided thoracic pedicle screw placement: a technical study in cadavers and preliminary clinical experience.

OBJECT: Thoracic pedicle screw fixation is effective and reliable in providing short-segment stabilization. Although the procedure is becoming more widely used, accurate insertion of the screws is difficult due to the small dimensions of thoracic pedicles, and the associated risk is high due to the proximity of the spinal cord. In previous studies authors have shown the accuracy of image-guided lumbar pedicle screw placement, but there have been no reported investigations into the accuracy of image-guided thoracic pedicle screw placement. The authors report their experience with such an investigation. METHODS: To evaluate the accuracy of image-guided thoracic pedicle screw placement in vitro and in vivo, thoracic pedicle screws were placed with an image-guidance system in five human cadavers and 10 patients. In cadavers, the accuracy of screw placement was assessed by postoperative computerized tomography and visual inspection and in patients by postoperative imaging studies. Of the 120 pedicle screws placed in five cadavers pedicle violation occurred in 23 cases (19.2%); there was one pedicle violation (4.2%) in each of the last two cadavers. Of the 45 pedicle screws placed in 10 patients, pedicle violations occurred in three (6.7%). CONCLUSIONS: In comparison with historical controls, the accuracy of thoracic pedicle screw placement is improved with the use of an image-guidance system. It allows the surgeon to visualize the thoracic pedicle and the surrounding structures that are normally out of the surgical field of view. The surgeon, however, must be aware of the limitations of an image-guidance system and have a sound basic knowledge of spinal anatomy to avoid causing serious complications.

Role of snare proteins in CFTR and ENaC trafficking.

The apical membrane ion channels, CFTR and ENaC, undergo regulated trafficking as a means of controlling their plasma membrane density. This provides a mechanism for regulating the Cl and Na conductance properties of epithelial apical membranes, and thus the transepithelial ion transport rates. Physical and functional interactions between these channels and SNARE proteins, in particular syntaxin 1A (S1A), provide a mechanism for linking the known vesicle fusion machinery with this process. In this paper we summarize evidence indicating that the interaction of S1A with CFTR and ENaC reduces channel currents in a syntaxin-isoform-specific manner. The acute cAMP-regulated CFTR trafficking event, which is reported by an increase in membrane capacitance in response to cAMP, is also inhibited by exogenous S1A expression. We tagged both channels with flag epitopes on their extracellular surfaces to monitor their plasma membrane expression as a function of S1A co-expression. The data indicate that the reduction in current caused by S1A is associated with a marked decrease in the amount of CFTR or ENaC detected at the cell surface. These findings suggest that S1A inhibits ion channel insertion into the plasma membrane, either by disrupting the stoichiometry of SNARE protein associations that mediate channel trafficking, or by physically associating with the channels to prevent their insertion. These data link the SNARE machinery to the regulation of apical membrane ion channel density, and suggest that phosphorylation-dependent interactions of these channels with SNARE proteins may acutely regulate this process.

Probing the interaction between inactivation gating and Dd-sotalol block of HERG.

Potassium channels encoded by HERG underlie I:(Kr), a sensitive target for most class III antiarrhythmic drugs, including methanesulfonanilides such as Dd-sotalol. Recently it was shown that these drugs are trapped in the channel as it closes during hyperpolarization. At the same time, HERG channels rapidly open and inactivate when depolarized, and methanesulfonanilide block is known to develop in a use-dependent manner, suggesting a potential role for inactivation in drug binding. However, the role of HERG inactivation in class III drug action is uncertain: pore mutations that remove inactivation reduce block, yet many of these mutations also modify the channel permeation properties and could alter drug affinity through gating-independent mechanisms. In the present study, we identify a definitive role for inactivation gating in Dd-sotalol block of HERG, using interventions complementary to mutagenesis. These interventions (addition of extracellular Cd(2+), removal of extracellular Na(+)) modify the voltage dependence of inactivation but not activation. In normal extracellular solutions, block of HERG current by 300 micromol/L Dd-sotalol reached 80% after a 10-minute period of repetitive depolarization to +20 mV. Maneuvers that impeded steady-state inactivation also reduced Dd-sotalol block of HERG: 100 micromol/L Cd(2+) reduced steady-state block to 55% at +20 mV (P:<0.05); removing extracellular Na(+) reduced block to 44% (P:<0.05). An inactivation-disabling mutation (G628C-S631C) reduced Dd-sotalol block to only 11% (P:<0.05 versus wild type). However, increasing the rate of channel inactivation by depolarizing to +60 mV reduced Dd-sotalol block to 49% (P:<0.05 versus +20 mV), suggesting that the drug does not primarily bind to the inactivated state. Coexpression of MiRP1 with HERG had no effect on inactivation gating and did not modify Dd-sotalol block. We postulate that Dd-sotalol accesses its receptor in the open pore, and the drug-receptor interaction is then stabilized by inactivation. Whereas deactivation traps the bound methanesulfonanilide during hyperpolarization, we propose that HERG inactivation stabilizes the drug-receptor interaction during membrane depolarization.

Non-enzymatic triggering of the ceramide signalling cascade by solar UVA radiation.

Ceramide is a key component of intracellular stress responses. Evidence is provided for a novel mechanism of ceramide formation that mediates solar ultraviolet (UV) A radiation-induced expression of the intercellular adhesion molecule (ICAM)-1. Similarly to UVA radiation, ceramide stimulation of human keratinocytes induced ICAM-1 mRNA expression and activated the ICAM-1 promoter through transcription factor AP-2. Ceramide-activated AP-2 and ceramide-induced ICAM-1 reporter gene activation were abrogated through deletion of the AP-2 binding site. UVA radiation increased the level of ceramide in keratinocytes and inhibition of sphingomyelin synthesis prevented UVA radiation-induced ICAM-1 expression. Hitherto, two pathways have been identified for ceramide accumulation: hydrolysis from sphingomyelin through neutral and acid sphingomyelinases, and de novo synthesis by ceramide synthase. UVA radiation did not activate any of these enzymes. Ceramide generation in UVA-irradiated cells, however, was inhibited by singlet oxygen quenchers and mimicked in unirradiated cells by a singlet oxygen-generating system. In addition, UVA radiation and singlet oxygen both generated ceramide in protein-free, sphingomyelin-containing liposomes. This study indicates that singlet oxygen triggers a third, non-enzymatic mechanism of ceramide formation.

Identification of the elements regulating the expression of the cell adhesion molecule MCAM/MUC18. Loss of AP-2 is not required for MCAM expression in melanoma cell lines.

The cell adhesion molecule melonoma cell adhesion molecule (MCAM)/MUC18/CD146 is specifically up-regulated on tumors of neuroectodermal origin and in animal models confers metastatic capacity to human melanoma cells. To identify critical regions regulating MCAM expression in melanomas, 1 kilobase of the MCAM 5' region was analyzed for promoter activity and transcription factor binding in 1 glioma, 1 carcinoma, and 4 melanoma cell lines. The minimal MCAM promoter (-106/+22 base pair (bp)) consists of 4 Sp-1 sites, two AP-2 elements, one cAMP responsive element, and the initiator surrounding the transcriptional start site. Analysis of mutated constructs indicated that the cAMP-responsive element is a major transcriptional activator in the majority of cell lines. Site-directed mutagenesis revealed that, in AP-2 expressing cells, the AP-2 site within the core promoter (-23 bp) has an inhibitory influence on MCAM expression while the AP-2 sites at -131 and -302 bp are activating. Functional AP-2 was observed in both MCAM positive and MCAM negative melanoma cell lines indicating that expression of MCAM does not require loss of this transcription factor. Furthermore, all MCAM constructs were strongly expressed in MCAM negative as well as MCAM positive cells, indicating that the expression of this gene is not controlled solely by the presence of transactivating factors binding to the investigated region.

Anterior cervical foraminotomy for unilateral radicular disease.

STUDY DESIGN: A clinical series of patients with unilateral radiculopathy treated with the anterior cervical foraminotomy procedure. OBJECTIVE: To establish procedural techniques and clinical and radiologic outcomes for the anterior cervical foraminotomy procedure. SUMMARY OF BACKGROUND DATA: Cervical radiculopathy is typically caused by unilateral disc herniation or uncovertebral osteophytes that compress the ventral aspect of the nerve. Direct removal of a cervical lesion causing radicular symptoms without concomitant fusion seems to be an ideal treatment in selected patients. The indications for an anterior cervical neural foraminotomy are limited to unilateral radicular symptoms at one or two levels, with minimal neck pain. METHODS: Twenty-one patients were treated with the anterior cervical neural foraminotomy procedure during a 3-year period with follow-up from 6 to 36 months. There were 13 men and 8 women (age range, 27-58 years). Fourteen patients had symptomatic soft disc herniation, and 7 had uncovertebral osteophytes confirmed by magnetic resonance imaging and/or myelogram and computed tomography. Sixteen patients had a single anterior cervical neural foraminotomy, and 5 had procedures at adjacent levels. RESULTS: Nineteen patients (91%) had improved or resolved radicular symptoms, and 2 (9%) had persistent radicular symptoms necessitating further surgery (one two-level anterior cervical neural discectomy and fusion and one posterior foraminal decompression). CONCLUSIONS: Patients treated with the anterior cervical neural foraminotomy procedure have equivalent or better outcomes than those who undergo current cervical procedures. It appears to be a good alternative procedure for carefully selected patients with unilateral cervical radiculopathy and avoids a fusion of the disc space.

Intracerebral clysis in a rat glioma model.

OBJECTIVE: Intracerebral clysis (ICC) is a new term we use to describe convection-enhanced microinfusion into the brain. This study establishes baseline parameters for preclinical, in vivo, drug investigations using ICC in a rat glioma model. METHODS: Intracranial pressure was measured, with an intraparenchymal fiber-optic catheter, in male Fischer rats 10, 15, 20, and 25 days after implantation of C6 glioma cells in the right frontal lobe (n = 80) and in control rats without tumor (n = 20), before and during ICC. A 25% albumin solution (100 microl) was infused through an intratumoral catheter at 0.5, 1.0, 2.0, 3.0, and 4.0 microl/min. Infusate distribution was assessed by infusion of fluorescein isothiocyanate-dextran (Mr 20,000), using the aforementioned parameters (n = 36). Brains were sectioned and photographed under ultraviolet light, and distribution was calculated by computer analysis (NIH Image for Macintosh). Safe effective drug distribution was demonstrated by measuring tumor sizes and apoptosis in animals treated with N,N'-bis(2-chloroethyl)-N-nitrosourea via ICC, compared with untreated controls. Magnetic resonance imaging noninvasively confirmed tumor growth before treatment. RESULTS: Intracranial pressure increased with tumor progression, from 5.5 mm Hg at baseline to 12.95 mm Hg on Day 25 after tumor cell implantation. Intracranial pressure during ICC ranged from 5 to 21 mm Hg and was correlated with increasing infusion volumes and increasing rates of infusion. No toxicity was observed, except at the higher ends of the tumor size and volume ranges. Fluorescein isothiocyanate-dextran distribution was greater with larger infusion volumes (30 microl versus 10 microl, n = 8, P < 0.05). No significant differences in distribution were observed when different infusion rates were compared while the volume was kept constant. At tolerated flow rates, the volumes of distribution were sufficient to promote adequate drug delivery to tumors. N,N'-Bis(2-chloroethyl)-N-nitrosourea treatment resulted in significant decreases in tumor size, compared with untreated controls. CONCLUSION: The C6 glioma model can be easily modified to study aspects of interstitial delivery via ICC and the application of ICC to the screening of potential antitumor agents for safety and efficacy.

Nephrotic syndrome in chronic lymphocytic leukemia: a paraneoplastic syndrome?

The association of malignancy and various glomerulopathies is a well recognized phenomenon. We report a case of nephrotic syndrome secondary to minimal-change disease in a patient with chronic lymphocytic leukemia (CLL) and review the literature on nephrotic syndrome in this disorder. Since the relative distribution of etiologies differ from what might be expected for primary nephrotic syndrome in a similar aged population, we propose that nephrotic syndrome is a potential paraneoplastic phenomenon associated with CLL. We hypothesize a possible etiologic role of different cell surface markers of the lymphocyte to explain the diverse renal histologic manifestations.

Treatment of posttraumatic syringomyelia with extradural decompressive surgery.

The authors review the management of five patients with posttraumatic syringomyelia (PTS) associated with an uncorrected spinal deformity. Patients with evidence of progressive neurological deterioration underwent ventral spinal decompressive surgery. The mean patient age at the time of injury was 39 years, and the time between injury and the diagnosis of PTS ranged from 2 to 22 years. Mechanisms of injury consisted of fracture/subluxations in three patients and burst fractures in two. All patients experienced delayed neurological deterioration consistent with PTS. Magnetic resonance imaging revealed ventral deformities, and the spinal canal stenosis ranged from 20 to 50% (mean 39%). All patients underwent ventral epidural spinal decompressive surgery to correct the bone deformity and restore the spinal canal. The mean follow-up period was 38 months. The decompressive intervention was initially successful in treating the neurological deterioration in all patients. Symptoms resolved completely in four patients, and the other experienced neurological improvement. Postoperative magnetic resonance imaging revealed a reduction in the size of syrinx cavity in the patients whose symptoms resolved and no change in the remaining patient. Two patients required a subsequent second-stage posterior intradural exploration and duraplasty for recurrence of symptoms and/or syrinx. Posttraumatic spinal deformity may cause spinal canal stenosis and alter subarachnoid cerebrospinal fluid (CSF) flow in certain patients. Ventral epidural spinal decompressive surgery may result in neurological improvement and a reduction of the syrinx cavity, avoiding the need for placement of a shunt or other intradural procedures. However, some patients will also require reconstruction of the posterior subarachnoid space with duraplasty if the ventral decompressive procedure achieves only partial restoration of the subarachnoid CSF flow.