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INTRODUCTION: The COVID-19 pandemic has put significant stress on healthcare systems globally. This study focuses on emergency general surgery services at a major trauma centre and teaching hospital. We aimed to identify whether the number of patients and the severity of their presentation has significantly changed since the implementation of a national lockdown. MATERIALS AND METHODS: This study is a retrospective review of acute referrals (from general practice and accident and emergency) to the emergency general surgery team over a 14-day period before (group 1) and during (group 2) lockdown. RESULTS: A total of 151 patients were reviewed by the general surgical team in group 1 and 75 in group 2 (a 50.3% reduction). The number of days with symptoms prior to presentation was significantly shorter in group 1 compared with group 2 (3 vs 4, p = 0.04). There was no significant difference in the National Early Warning Score, white blood cell count, lymphocytes and C-reactive protein on admission between the two groups of patients. There were significantly fewer patients admitted after lockdown compared with pre-lockdown (66% vs 48%, p = 0.01). Length of hospital stay was significantly shorter during lockdown compared with pre-lockdown (5 days vs 4 days, p = 0.04). CONCLUSION: Fewer patients were referred and admitted during lockdown compared with pre-lockdown, and the length of stay was also significantly reduced. There was also a delay in presentation to hospital, although these patients were not more unwell based on the scoring criteria used within this study.
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AIMS: The spatial resolution of light microscopy is limited by the wavelength of visible light (the 'diffraction limit', approximately 250 nm). Resolution of sub-cellular structures, smaller than this limit, is possible with super resolution methods such as stochastic optical reconstruction microscopy (STORM) and super-resolution optical fluctuation imaging (SOFI). We aimed to resolve subcellular structures (axons, myelin sheaths and astrocytic processes) within intact white matter, using STORM and SOFI. METHODS: Standard cryostat-cut sections of subcortical white matter from donated human brain tissue and from adult rat and mouse brain were labelled, using standard immunohistochemical markers (neurofilament-H, myelin-associated glycoprotein, glial fibrillary acidic protein, GFAP). Image sequences were processed for STORM (effective pixel size 8-32 nm) and for SOFI (effective pixel size 80 nm). RESULTS: In human, rat and mouse, subcortical white matter high-quality images for axonal neurofilaments, myelin sheaths and filamentous astrocytic processes were obtained. In quantitative measurements, STORM consistently underestimated width of axons and astrocyte processes (compared with electron microscopy measurements). SOFI provided more accurate width measurements, though with somewhat lower spatial resolution than STORM. CONCLUSIONS: Super resolution imaging of intact cryo-cut human brain tissue is feasible. For quantitation, STORM can under-estimate diameters of thin fluorescent objects. SOFI is more robust. The greatest limitation for super-resolution imaging in brain sections is imposed by sample preparation. We anticipate that improved strategies to reduce autofluorescence and to enhance fluorophore performance will enable rapid expansion of this approach.
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Encéfalo/diagnóstico por imagen , Microscopía/métodos , Sustancia Blanca/diagnóstico por imagen , Anciano , Anciano de 80 o más Años , Animales , Femenino , Humanos , Procesamiento de Imagen Asistido por Computador , Masculino , Ratones , Persona de Mediana Edad , RatasRESUMEN
AIM: Imaging for pelvic floor defaecatory dysfunction includes defaecation proctography. Integrated total pelvic floor ultrasound (transvaginal, transperineal, endoanal) may be an alternative. This study assesses ultrasound accuracy for the detection of rectocele, intussusception, enterocele and dyssynergy compared with defaecation proctography, and determines if ultrasound can predict symptoms and findings on proctography. Treatment is examined. METHOD: Images of 323 women who underwent integrated total pelvic floor ultrasound and defaecation proctography between 2011 and 2014 were blindly reviewed. The size and grade of rectocele, enterocele, intussusception and dyssynergy were noted on both, using proctography as the gold standard. Barium trapping in a rectocele or a functionally significant enterocele was noted on proctography. Demographics and Obstructive Defaecation Symptom scores were collated. RESULTS: The positive predictive value of ultrasound was 73% for rectocele, 79% for intussusception and 91% for enterocele. The negative predictive value for dyssynergy was 99%. Agreement was moderate for rectocele and intussusception, good for enterocele and fair for dyssynergy. The majority of rectoceles that required surgery (59/61) and caused barium trapping (85/89) were detected on ultrasound. A rectocele seen on both transvaginal and transperineal scanning was more likely to require surgery than if seen with only one mode (P = 0.0001). If there was intussusception on ultrasound the patient was more likely to have surgery (P = 0.03). An enterocele visualized on ultrasound was likely to be functionally significant on proctography (P = 0.02). There was, however, no association between findings on imaging and symptoms. CONCLUSION: Integrated total pelvic floor ultrasound provides a useful screening tool for women with defaecatory dysfunction such that defaecatory imaging can avoided in some.
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Estreñimiento/diagnóstico por imagen , Defecografía/métodos , Endosonografía/métodos , Trastornos del Suelo Pélvico/diagnóstico por imagen , Diafragma Pélvico/diagnóstico por imagen , Adulto , Anciano , Anciano de 80 o más Años , Ataxia/complicaciones , Ataxia/diagnóstico por imagen , Ataxia/fisiopatología , Bario , Estreñimiento/etiología , Estreñimiento/fisiopatología , Medios de Contraste , Defecación/fisiología , Femenino , Hernia/complicaciones , Hernia/diagnóstico por imagen , Hernia/fisiopatología , Humanos , Intususcepción/complicaciones , Intususcepción/diagnóstico por imagen , Intususcepción/fisiopatología , Persona de Mediana Edad , Diafragma Pélvico/fisiopatología , Trastornos del Suelo Pélvico/complicaciones , Trastornos del Suelo Pélvico/fisiopatología , Valor Predictivo de las Pruebas , Rectocele/complicaciones , Rectocele/diagnóstico por imagen , Rectocele/fisiopatología , Índice de Severidad de la Enfermedad , Método Simple CiegoRESUMEN
AIM: The study aimed to determine the current state of UK pelvic floor services and to discuss future strategies. METHOD: A questionnaire developed by the Pelvic Floor Society was sent in 2014 to the 175 colorectal units recognized by the Association of Coloproctology of Great Britain and Ireland. Questions included type of centre, frequency of pelvic floor clinics/interdisciplinary joint pelvic floor clinics/multidisciplinary meetings (MDMs) and workload. RESULTS: Sixty-seven (38%) centres replied including 75% of units with a consultant who was as member of the Pelvic Floor Society. Of the 67 centres 39% were tertiary centres for pelvic floor surgery (tertiary), 48% performed some pelvic floor surgery (regional) and 13% did not perform any (local). Ninety-six per cent of tertiary referral centres served a population over 500 000. The mean number of whole time equivalent consultants in tertiary centres was 1.03 and 0.77 in regional centres. Eighty per cent of tertiary centres and 56% of regional centres ran pelvic floor clinics. Eighty-four per cent of tertiary referral and 75% of regional units held or attended an MDM. Anal ultrasonography, anorectal physiology and proctography were performed in 96% of tertiary centres compared with 50% of non-tertiary units. CONCLUSION: The provision of pelvic floor services includes local, regional and tertiary centres. The overall response rate was low (38%) and biased to centres with a consultant who was a member of the Pelvic Floor Society. Not all regional or tertiary centres held an MDM or a pelvic floor clinic. Given the nature of pelvic floor pathology an integrated service should be aimed at linking different centres and specialities.
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Cirugía Colorrectal/tendencias , Predicción , Encuestas de Atención de la Salud , Trastornos del Suelo Pélvico , Cirugía Colorrectal/métodos , Humanos , Irlanda , Grupo de Atención al Paciente/tendencias , Encuestas y Cuestionarios , Reino UnidoRESUMEN
Carotid artery disease is a widespread cause of morbidity and mortality. Porcine models of vascular disease are well established in vivo, but existing endothelial systems in vitro (e.g. human umbilical vein endothelial cells, rat aortic endothelial cultures) poorly reflect carotid endothelium. A reliable in vitro assay would improve design of in vivo experiments and allow reduction and refinement of animal use. This study aimed (1) to develop ex vivo endothelial cultures from porcine carotid and (2) to test whether these were suitable for lentivector-mediated transgene delivery. Surplus carotid arteries were harvested from young adult female Large White pigs within 10 min post-mortem. Small sectors of carotid artery wall (approximately 4 mm×4 mm squares) were immobilised in a stable gel matrix. Cultures were exposed to HIV-derived lentivector (LV) encoding a reporter transgene or the equivalent integration-deficient vector (IDLV). After 7-14 days in vitro, cultures were fixed and labelled histochemically. Thread-like multicellular outgrowths were observed that were positive for endothelial cell markers (CD31, VEGFR2, von Willebrand factor). A minority of cells co-labelled for smooth muscle markers. Sensitivity to cytotoxic agents (paclitaxel, cycloheximide, staurosporine) was comparable to that in cell cultures, indicating that the gel matrix permits diffusive access of small pharmacological molecules. Transgene-expressing cells were more abundant following exposure to LV than IDLV (4.7, 0.1% of cells, respectively). In conclusion, ex vivo adult porcine carotid artery produced endothelial cell outgrowths that were effectively transduced by LV. This system will facilitate translation of novel therapies to clinical trials, with reduction and refinement of in vivo experiments.
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Arterias Carótidas/citología , Endotelio Vascular/citología , Vectores Genéticos , Lentivirus/genética , Animales , Femenino , Técnicas de Transferencia de Gen , Porcinos , TransgenesRESUMEN
The principles of fast track surgery are well established in colorectal surgery. It is an evidence based model aimed at reducing length of stay, convalescence and morbidity by optimising both clinical and organisational factors. Despite this, the implementation of fast track surgery in breast cancer patients has been slow. The 23 h discharge model for breast cancer surgery patients has been outlined by the NHS Improvement Programme and is a breakthrough from traditional inpatient care. This paper outlines the early experience of implementation of this model in a single institution during a 3-month audit period. Over 80% of patients undergoing non-reconstructive breast surgery were discharged safely within '23 h'. This suggests that good communication links, reorganisation of existing resources, active user involvement (both patients and clinical team) and strong project management ensures fast-tracking to be safe for the patient with significant economic benefits for the hospital.
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Neoplasias de la Mama/economía , Neoplasias de la Mama/cirugía , Costos de Hospital , Tiempo de Internación , Mastectomía/métodos , Adulto , Anciano , Anciano de 80 o más Años , Protocolos Clínicos , Drenaje , Femenino , Humanos , Tiempo de Internación/economía , Mastectomía/efectos adversos , Mastectomía/economía , Persona de Mediana Edad , Alta del Paciente , Factores de Tiempo , Adulto JovenRESUMEN
AIMS: Serum- and glucocorticoid-inducible kinase 1 (SGK1) protects neuronal cells from injury stimuli in vitro, and exerts anti-apoptotic effects via downstream targets including the forkhead-like transcription factor FOXO3a. SGK1 is a homolog of Akt, a related survival kinase that is up-regulated in Alzheimer's disease (AD). Here we aimed to examine the expression pattern of SGK1 and FOXO3a in aged human cerebral cortex. METHODS: Cortical tissue from aged donors without brain disease (aged controls, AC, n = 19) and from severe AD patients (Braak stage V-VI; n = 14) were examined by immunohistochemistry and immunoblot analysis. RESULTS: SGK1 was present in all samples (detected by immunohistochemistry and immunoblotting). Large cortical neuronal cells were strongly positive for SGK1, with predominantly nuclear labelling. Some astrocytes and oligodendrocytes were also labelled. SGK1 was not seen in nerve tracts (axons or myelin) and rarely seen in CD68-positive cells (microglia, perivascular macrophages) or vascular cells (myocytes or endothelia). The fraction of large cortical neurones with nuclear FOXO3a was lower in AD cases relative to AC (54%, 70%, respectively, P < 0.001). In immunoblots no difference in SGK1 abundance was detected between AC and AD tissues. Phosphorylation of NDRG1 (an SGK1-specific target) was greater in AD, relative to AC cases (approximately twofold, P = 0.023). CONCLUSIONS: Neuronal expression of SGK1 in aged human brain and its nuclear compartmentalization suggest a possible neuroprotective role. FOXO3a and NDRG1 data suggest augmented SGK1 activity (as reported for Akt) in severe AD. Increased intracellular SGK1 may complement enhanced Akt, with a distinct subcellular localization.
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Envejecimiento/metabolismo , Enfermedad de Alzheimer/metabolismo , Corteza Cerebral/metabolismo , Factores de Transcripción Forkhead/metabolismo , Proteínas Inmediatas-Precoces/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Anciano , Anciano de 80 o más Años , Enfermedad de Alzheimer/enzimología , Proteínas de Ciclo Celular/metabolismo , Supervivencia Celular , Corteza Cerebral/citología , Corteza Cerebral/enzimología , Femenino , Proteína Forkhead Box O3 , Humanos , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Masculino , Neuronas/metabolismoRESUMEN
There are few published data on aldosterone and cortisol co-secreting adrenal tumours. Failure to perform comprehensive preoperative endocrine investigations in patients with adrenal "incidentalomas" or in those thought to be secreting only one hormone may account for this. Clinically patients with such lesions may have evidence of hypertension and hypokalaemia with no features of cortisol excess. Preoperative diagnosis of such lesions with accurate endocrinological work up is essential to prevent adrenal insufficiency and haemodynamic crises following removal of such glands. We present a series of 4 patients with co-secreting tumours treated by laparoscopic adrenalectomy between September 2010 and March 2011. Our experience suggests that dual secretors are more common than originally thought. A high index of suspicion and adequate endocrine work up is paramount in diagnosing such tumours and in experienced hands, laparoscopic adrenalectomy with appropriate substitutive steroid cover is safe, feasible and curative for these functioning adrenal tumours.
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Adenoma/metabolismo , Neoplasias de las Glándulas Suprarrenales/metabolismo , Neoplasias de las Glándulas Suprarrenales/cirugía , Adrenalectomía/métodos , Aldosterona/metabolismo , Hidrocortisona/metabolismo , Laparoscopía/métodos , Adenoma/diagnóstico , Adenoma/cirugía , Neoplasias de las Glándulas Suprarrenales/diagnóstico , Anciano , Anciano de 80 o más Años , Aldosterona/sangre , Femenino , Humanos , Hidrocortisona/sangre , Masculino , Atención Perioperativa , Tomografía Computarizada por Rayos XRESUMEN
BACKGROUND: Perineal wound complications following abdominoperineal excision (APE) for low rectal tumours remain an important cause of morbidity and prolonged hospital stay, particularly after chemoradiotherapy. The aim was to assess outcomes after using inferior gluteal artery perforator (IGAP) flaps for immediate perineal reconstruction, and to compare these with the authors' previous experience and published literature on myocutaneous flaps. METHODS: A series of patients who underwent immediate IGAP flap reconstruction after APE between April 2008 and December 2010 were examined retrospectively to determine patient demographics, length of operation, complications (perineal wound and general) and length of hospital stay. RESULTS: Forty patients with rectal adenocarcinoma (33 primary and 7 recurrent disease) underwent immediate IGAP flap reconstruction following APE. Median follow-up was 9 months. Neoadjuvant chemoradiotherapy was received by 98 per cent of the patients. Thirty-two patients underwent APE plus IGAP flaps (25 open, 7 laparoscopic), with a median operating time of 402 min, and eight patients had multivisceral resection (MVR) plus IGAP flaps (7 total pelvic exenteration (TPE), 1 abdominosacral resection), with a median duration of surgery of 561 min. There was one death (fatal stroke) and four major flap complications (10 per cent) (1 enteroperineal fistula, and 3 deep wound infections). Median length of hospital stay was 13 days after APE plus IGAP flaps and 27 days following MVR plus IGAP flaps. Late complications occurred in two patients who had vaginal reconstruction and developed perineal hernias requiring revisional surgery. CONCLUSION: Although operating times are long, the IGAP flap is robust, with no flap necrosis observed in this series.
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Adenocarcinoma/cirugía , Perineo/cirugía , Neoplasias del Recto/cirugía , Colgajos Quirúrgicos/irrigación sanguínea , Adulto , Anciano , Nalgas/irrigación sanguínea , Nalgas/cirugía , Quimioradioterapia Adyuvante/métodos , Femenino , Humanos , Tiempo de Internación/estadística & datos numéricos , Masculino , Persona de Mediana Edad , Recurrencia Local de Neoplasia , Colgajos Quirúrgicos/efectos adversos , Resultado del TratamientoRESUMEN
OBJECTIVES: Cerebral small vessel disease (SVD) is common in aged brains and causes lacunar stroke, diffuse white matter lesions (leukoaraiosis), and vascular cognitive impairment. The pathogenesis is unknown. Endothelial dysfunction is a possible causal factor, and circulating markers of endothelial activation (intercellular adhesion molecule-1, thrombomodulin) and inflammation (interleukin [IL]-6) are elevated in patients with SVD. In this case-control study, we tested whether brain endothelial ICAM1, thrombomodulin, and IL-6 are altered in SVD. METHODS: We examined small penetrating cerebral arteries of pathologically diagnosed SVD cases, aged controls without SVD, young control cases with no brain pathology, and cases with early-onset hereditary SVD (cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy [CADASIL]). All tissues had minimal cerebral amyloid angiopathy or other Alzheimer pathology. RESULTS: Thrombomodulin immunoreactivity was present in all aged SVD, aged control, and CADASIL cases, primarily in small artery endothelium. Thrombomodulin was augmented in aged SVD cases compared with aged controls (p = 0.012) and in vessels with higher sclerotic index (an indicator of SVD severity; p < 0.01). Thrombomodulin was sparse/absent in young controls. Endothelial ICAM1 and IL-6 were rarely seen, and were not related to SVD. CONCLUSIONS: Our data suggest that cerebral endothelial activation in deep penetrating arteries is not associated with SVD. Endothelial thrombomodulin increased with SVD severity, and CADASIL data suggest that this may be a cerebral response to SVD. Elevated thrombomodulin may be a protective agent in SVD. Our data confirm endothelial involvement in SVD.
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Envejecimiento/metabolismo , Envejecimiento/patología , Enfermedades de los Pequeños Vasos Cerebrales/metabolismo , Enfermedades de los Pequeños Vasos Cerebrales/patología , Endotelio Vascular/metabolismo , Endotelio Vascular/patología , Trombomodulina/metabolismo , Anciano , Anciano de 80 o más Años , Femenino , Humanos , MasculinoAsunto(s)
Carcinoma de Células Escamosas/diagnóstico , Reservorios Cólicos/patología , Neoplasias del Íleon/diagnóstico , Carcinoma de Células Escamosas/complicaciones , Carcinoma de Células Escamosas/cirugía , Femenino , Humanos , Histerectomía , Neoplasias del Íleon/complicaciones , Neoplasias del Íleon/cirugía , Persona de Mediana Edad , Invasividad Neoplásica/diagnóstico , Ovariectomía , Reservoritis/etiología , Reservoritis/patología , SalpingectomíaRESUMEN
BACKGROUND: Transient receptor potential vanilloid subtype 1 (TRPV1) receptor is a primary pain-sensing relay at peripheral sensory nerve endings and is also widespread in the brain, where it is implicated in neurodegeneration. Previous studies of TRPV1 neurotoxicity have utilized heterogeneous receptor populations, non-selective ligands, or non-neuronal cell types. Here, we explored the pharmacology of TRPV1-induced cytotoxicity in a homogeneous, neurone-like cellular environment. METHODS: Cell death was examined in a human neurone-like cell line, stably expressing recombinant human TRPV1. Cytotoxicity was quantified in terms of nuclear morphology and mitochondrial complex II activity. Immunocytochemical markers of apoptotic cell death were also examined. RESULTS: The TRPV1-selective agonist capsaicin, and the endovanilloids anandamide and N-arachidonoyl-dopamine (NADA), induced TRPV1-dependent delayed cell death in a concentration- and time-dependent manner. Capsaicin exposure time was significantly correlated with potency (r(2)=0.91, P=0.01). Release of cytochrome c from mitochondria, activation of caspase-3, and condensed nuclear chromatin were evident 6 h after capsaicin exposure, but cytotoxicity was unaffected by a pan-caspase inhibitor (zVAD-fmk, 50 microM). CONCLUSIONS: We conclude that capsaicin, anandamide, and NADA can initiate TRPV1-dependent delayed cell death in neurone-like cells. This is an apoptosis-like process, but independent of caspase activity.
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Apoptosis/efectos de los fármacos , Ácidos Araquidónicos/farmacología , Capsaicina/farmacología , Dopamina/análogos & derivados , Neuronas/efectos de los fármacos , Alcamidas Poliinsaturadas/farmacología , Canales Catiónicos TRPV/fisiología , Dopamina/farmacología , Relación Dosis-Respuesta a Droga , Endocannabinoides , Humanos , Neuronas/metabolismo , Neuronas/patología , Canales Catiónicos TRPV/agonistas , Canales Catiónicos TRPV/metabolismo , Células Tumorales CultivadasRESUMEN
AIMS: Here our objective was to detect the pro-apoptotic serine/threonine kinase death-associated protein kinase (DAPK1) in aged human cerebral cortex and to test the hypothesis that DAPK1 abundance is associated with late-onset Alzheimer's disease (AD). METHODS: Using Western analysis and immunohistochemistry we evaluated post mortem frontal cerebral cortex from patients with severe AD (mean age 76 years, range 66-91, n = 11, all male), and from control cases without serious central nervous system illness (mean age 77 years, range 61-95, n = 12, all male). We also examined brains of Tg2576 transgenic mice (males, aged 16-21 months), a model for chronic amyloid-induced brain injury. RESULTS: Immunohistochemical labelling showed DAPK1 expression in cortical neurones of human cortex and axonal tracts within subcortical white matter, both in AD and in control brains. Western analysis confirmed DAPK1 expression in all samples, although expression was very low in some control cases. DAPK1 abundance in the AD group was not significantly different from that in controls (P = 0.07, Mann-Whitney test). In brains of Tg2576 mice DAPK1 abundance was very similar to that in wild-type littermates (P = 0.96, Mann-Whitney test). CONCLUSION: We found that DAPK1 was expressed in neurones of aged human frontal cortex, both in AD and in control cases.
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Enfermedad de Alzheimer/enzimología , Proteínas Reguladoras de la Apoptosis/biosíntesis , Proteínas Quinasas Dependientes de Calcio-Calmodulina/biosíntesis , Corteza Cerebral/enzimología , Edad de Inicio , Anciano , Anciano de 80 o más Años , Enfermedad de Alzheimer/patología , Animales , Western Blotting , Corteza Cerebral/patología , Proteínas Quinasas Asociadas a Muerte Celular , Humanos , Inmunohistoquímica , Masculino , Ratones , Ratones TransgénicosRESUMEN
Deep brain stimulation (DBS) of the subthalamic nucleus (STN) in Parkinson's disease (PD) patients augments STN-driven excitation of the internal globus pallidus (GPi). However, other DBS-induced changes are largely unknown. Here we report the biochemical effects of STN-DBS in two basal ganglia stations (putamen--PUT--and GPi) and in a thalamic relay nucleus, the anteroventral thalamus (VA). In six advanced PD patients undergoing surgery, microdialysis samples were collected from GPi, PUT and VA before, during and after one hour of STN-DBS. cGMP was measured in the GPi and PUT as an index of glutamatergic transmission, whereas GABA was measured in the VA. During clinically effective STN-DBS, we found a significant decrease in GABA extracellular concentrations in the VA (-25%). Simultaneously, cGMP extracellular concentrations were enhanced in the PUT (+200%) and GPi (+481%). DBS differentially affects fibers crossing the STN area: it activates the STN-GPi pathway while inhibiting the GPi-VA one. These findings support a thalamic dis-inhibition, as the main responsible for the clinical effect of STN-DBS. This, in turn, re-establishes a more physiological level of PUT activity.
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Estimulación Encefálica Profunda , Enfermedad de Parkinson/metabolismo , Enfermedad de Parkinson/terapia , Anciano , Biomarcadores , GMP Cíclico/metabolismo , Espacio Extracelular/metabolismo , Femenino , Globo Pálido/metabolismo , Humanos , Masculino , Microdiálisis , Persona de Mediana Edad , Tálamo/metabolismo , Ácido gamma-Aminobutírico/metabolismoRESUMEN
202W92 (R-(-)-2,4-diamino-6-(fluromethyl)-5-(2,3,5-trichlorophenyl)pyrimidine) is a novel compound in the same chemical series as the antiepileptic drug lamotrigine and the neuroprotective sipatrigine. Here 202W92 was quantitatively assessed as a neuroprotective agent in focal cerebral ischaemia, and as an inhibitor of sodium and calcium channels and of synaptic transmission. In the rat permanent middle cerebral artery occlusion (MCAO) model of acute focal ischaemia, 202W92 reduced infarct volume by 75% in cortex and by 80% in basal ganglia, with ED(50) approximately 2 mg/kg (single i.v. dose, 10 min post-occlusion). In whole-cell current recordings from single cells, 202W92 completely and reversibly inhibited voltage gated sodium channels (IC(50) 3 x 10(-6) M) in rat freshly-isolated cortical neurons and in the GH(3) pituitary cell line. 202W92 also inhibited a nifedipine-sensitive fraction (approximately 35%) of native high-voltage-activated (HVA) calcium current in rat cortical neurons (IC(50) 15 x 10(-6) M) and weakly inhibited low-voltage-activated (LVA) calcium currents of the recombinant alpha1I-mediated T-type (IC(50)>100 x 10(-6) M). The drug inhibited the amplitude and frequency of 4-aminopyridine-evoked glutamatergic excitatory post-synaptic currents (EPSCs). In conclusion, 202W92 is an effective neuroprotective agent when administered post-ischaemia and a potent sodium channel inhibitor in vitro.
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Isquemia Encefálica/tratamiento farmacológico , Canales de Calcio/efectos de los fármacos , Fármacos Neuroprotectores/farmacología , Pirimidinas/farmacología , Bloqueadores de los Canales de Sodio , Transmisión Sináptica/efectos de los fármacos , Telencéfalo/efectos de los fármacos , Animales , Isquemia Encefálica/metabolismo , Isquemia Encefálica/fisiopatología , Bloqueadores de los Canales de Calcio/farmacología , Canales de Calcio/metabolismo , Células Cultivadas/efectos de los fármacos , Células Cultivadas/metabolismo , Infarto Cerebral/tratamiento farmacológico , Infarto Cerebral/metabolismo , Infarto Cerebral/fisiopatología , Relación Dosis-Respuesta a Droga , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Potenciales Postsinápticos Excitadores/fisiología , Ácido Glutámico/metabolismo , Infarto de la Arteria Cerebral Media/tratamiento farmacológico , Infarto de la Arteria Cerebral Media/metabolismo , Infarto de la Arteria Cerebral Media/fisiopatología , Masculino , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/fisiología , Degeneración Nerviosa/tratamiento farmacológico , Degeneración Nerviosa/metabolismo , Degeneración Nerviosa/fisiopatología , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Técnicas de Placa-Clamp , Ratas , Ratas Endogámicas F344 , Canales de Sodio/metabolismo , Sustancia Negra/efectos de los fármacos , Sustancia Negra/metabolismo , Sustancia Negra/fisiopatología , Transmisión Sináptica/fisiología , Telencéfalo/metabolismo , Telencéfalo/fisiopatologíaRESUMEN
We have investigated the neuroprotective properties of AR-A008055 [(+/-)-1-(4-methyl-5-thiazolyl-1-phenyl-methylamine], a novel compound structurally related to clomethiazole. Administration (i.p.) of (+/-)-AR-A008055 60 min after 5 min of global cerebral ischaemia in gerbils produced a dose-dependent protection of the hippocampus from damage. Both enantiomers [(R)-(+)-AR-A008055 and (S)-(-)- AR-A008055] at 600 micromol/kg produced similar protection to that following clomethiazole (600& micromol/kg) and both produced similar and sustained neuroprotection, at 4, 7 and 21 days post-insult. When infused intravenously over a 2-h period, both enantiomers produced concentration-dependent neuroprotection, with the enantiomers providing similar protection at every plasma concentration (50-200 nmol/ml). The efficacy of (S)-(-)-AR-A008055 was similar to clomethiazole, but it was slightly less potent. Ischaemia-induced glutamate efflux from rat brain cortical prisms in vitro was inhibited by both isomers (100 microM). The inhibitory effect of (R)-(+)-AR-A008055 was blocked by bicuculline (10 microM) and picrotoxin (100 microM), while the effect of (S)-(-)-AR-A008055 was only antagonised by picrotoxin. This indicated that (S)-(-)-AR-A008055, like clomethiazole, is able to open the GABA(A)-chloride channel in the absence of endogenous GABA. (R)-(+)-AR-A008055 was more potent than (S)-(-)-AR-A008055 in enhancing the concentration of GABA in the medium following 30 min exposure of tissue to the ischaemic conditions, suggesting that it is an effective GABA uptake inhibitor. This action may explain both its effect on glutamate efflux in vitro and its neuroprotective effect in vivo.
Asunto(s)
Isquemia Encefálica/metabolismo , Clormetiazol/análogos & derivados , Clormetiazol/farmacología , Ácido Glutámico/metabolismo , Hipocampo/efectos de los fármacos , Metilaminas/farmacología , Fármacos Neuroprotectores/farmacología , Tiazoles/farmacología , Ácido gamma-Aminobutírico/metabolismo , Animales , Femenino , Gerbillinae , Hipocampo/metabolismo , Masculino , Fármacos Neuroprotectores/química , Ratas , Ratas Wistar , Accidente Cerebrovascular/metabolismoRESUMEN
AR-A008055 [(+/-)-1-(4-methyl-5-thiazolyl)-1-phenylmethylamine] is structurally related to clomethiazole and has been used to probe the mechanism of the neuroprotective effect of clomethiazole. Clomethiazole, (+/-)-AR-A008055 and (S)-(-)-AR-A008055 all displaced [35S]-t-butyl-bicyclophosphorothionate ([35S]TBPS) from rat cerebral cortex tissue (IC50 values: GABA, 8.1+/-0.04 microM; clomethiazole, 130+/-30 microM; (+/-)-AR-A008055, 494+/-7 microM; (S)-(-)-AR-A008055, 221+/-14 microM. (R)-(+)-AR-A008055 was without significant effect (IC50>1000 microM). None of the compounds interacted with NMDA or AMPA receptors or with sodium or calcium (N, P/Q) channels. Brain penetration of both enantiomers following their i.p. administration was excellent, with brain and plasma concentrations being similar. Clomethiazole dose-dependently inhibited spontaneous locomotor activity in rats and was approximately 10 times more sedative than either enantiomer of AR-A008055. Clomethiazole was more potent than (S)-(-)-AR-A008055 in the "pull-up" test (muscle relaxation) and in producing loss of righting reflex, while (R)-(+)-AR-A008055 had little effect. The time animals remained on a Rota-rod was of the order: clomethiazole<(S)-(-)-AR-A008055<(R)-(+)-AR-A008055. (S)-(-)-AR-A008055 (210 micromol/kg) raised seizure threshold to pentylenetetrazole (i.v.) by 119+/-21%. The (R)-(+)- enantiomer was not anticonvulsant. Overall, (S)-(-)-AR-A008055 exhibited a similar pharmacology to clomethiazole. However, its sedative and muscle relaxant effects were substantially less than clomethiazole, emphasising that these properties are not directly related to neuroprotective efficacy. The current data suggest that the proposed GABA uptake inhibitory property of (R)-(+)-AR-A008055 fails to produce significant sedative, myorelaxant or anticonvulsant activity.
Asunto(s)
Encéfalo/efectos de los fármacos , Clormetiazol/farmacología , Moduladores del GABA/farmacología , Metilaminas/farmacología , Actividad Motora/efectos de los fármacos , Relajación Muscular/efectos de los fármacos , Receptores de GABA-A/metabolismo , Tiazoles/farmacología , Animales , Anticonvulsivantes/química , Anticonvulsivantes/farmacología , Encéfalo/metabolismo , Compuestos Bicíclicos Heterocíclicos con Puentes/metabolismo , Clormetiazol/análogos & derivados , Clormetiazol/sangre , Convulsivantes/metabolismo , Relación Dosis-Respuesta a Droga , Moduladores del GABA/sangre , Moduladores del GABA/química , Hipnóticos y Sedantes/química , Hipnóticos y Sedantes/farmacología , Masculino , Actividad Motora/fisiología , Relajantes Musculares Centrales/química , Relajantes Musculares Centrales/farmacología , Relajación Muscular/fisiología , RatasRESUMEN
Acidic extracellular pH reduced high-voltage-activated (HVA) currents in freshly isolated cortical pyramidal neurones of adult rats, shifting activation to more positive voltages (V(1/2)=-18 mV at pH 7.4, -11 mV at pH 6.4). Sipatrigine inhibited HVA currents, with decreasing potency at acidic pH (IC(50) 8 microM at pH 7.4, 19 microM at pH 6.4) but the degree of maximal inhibition was >80% in all cases (pH 6.4-8.0). Sipatrigine has two basic groups (pK(A) values 4.2, 7.7) and at pH 7.4 is 68% in monovalent cationic form and 32% uncharged. From simple binding theory, the pH dependence of sipatrigine inhibition indicates a protonated group with pK(A) 6.6. Sipatrigine (50 microM) shifted the voltage dependence of channel activation at pH 7.4 (-7.6 mV shift) but not at pH 6.4. Lamotrigine has one basic site (pK(A) 5.5) and inhibited 34% of the HVA current, with similar potency over the pH range 6.4--7.4 (IC(50) 7.5--9 microM). These data suggest that the sipatrigine binding site on HVA calcium channels binds both cationic and neutral forms of sipatrigine, interacts with a group with pK(A)=6.6 and with the channel activation process, and differs from that for lamotrigine.