Shared functional defect in IP3R-mediated calcium signaling in diverse monogenic autism syndromes.

Autism spectrum disorder (ASD) affects 2% of children, and is characterized by impaired social and communication skills together with repetitive, stereotypic behavior. The pathophysiology of ASD is complex due to genetic and environmental heterogeneity, complicating the development of therapies and making diagnosis challenging. Growing genetic evidence supports a role of disrupted Ca(2+) signaling in ASD. Here, we report that patient-derived fibroblasts from three monogenic models of ASD-fragile X and tuberous sclerosis TSC1 and TSC2 syndromes-display depressed Ca(2+) release through inositol trisphosphate receptors (IP3Rs). This was apparent in Ca(2+) signals evoked by G protein-coupled receptors and by photoreleased IP3 at the levels of both global and local elementary Ca(2+) events, suggesting fundamental defects in IP3R channel activity in ASD. Given the ubiquitous involvement of IP3R-mediated Ca(2+) signaling in neuronal excitability, synaptic plasticity, gene expression and neurodevelopment, we propose dysregulated IP3R signaling as a nexus where genes altered in ASD converge to exert their deleterious effect. These findings highlight potential pharmaceutical targets, and identify Ca(2+) screening in skin fibroblasts as a promising technique for early detection of individuals susceptible to ASD.

An unusual intracranial tumour presenting in pregnancy.

We describe a patient who presented in late pregnancy with deteriorating neurological status due to an intracranial capillary haemangioma causing mass effect and raised intracranial pressure. She became confused and uncooperative leading to practical difficulties in performing adequate radiological imaging. Decision regarding timing of delivery and craniotomy was not straightforward and required discussion between the neurosurgeon, obstetrician and anaesthetist based on assessment of fetal maturity and the need to perform a craniotomy to excise what was initially thought to be a meningioma. Caesarean section was performed under general anaesthesia. The tumour was resected three weeks later. Management of obstetric patients with brain tumours is complex, requiring knowledge of the physiological effects of pregnancy on tumour size and labour on intracranial pressure. Both of these may influence the choice of labour analgesia or anaesthesia for caesarean section. Anaesthetists must be aware of the difficulties of radiological imaging during pregnancy, particularly in confused patients. The conflicting requirements of general anaesthesia for craniotomy and caesarean section should be considered.

Role of calcium in the pathogenesis of Alzheimer's disease and transgenic models.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder of the elderly that is characterized by memory loss. Neuropathologically, the AD brain is marked by an increased AP burden, hyperphosphorylated tau aggregates, synaptic loss, and inflammatory responses. Disturbances in calcium homeostasis are also one of the earliest molecular changes that occur in AD patients, alongside alterations in calcium-dependent enzymes in the post-mortem brain. The sum of these studies suggests that calcium dyshomeostasis is an integral part of the pathology, either influencing AP production, mediating its effects or both. Increasing evidence from in vitro studies demonstrates that the AP peptide could modulate a number of ion channels increasing calcium influx, including voltage-gated calcium and potassium channels, the NMDA receptor, the nicotinic receptor, as well as forming its own calcium-conducting pores. In vivo evidence has shown that A3 impairs both LTP and cognition, whereas all of these ion channels cluster at the synapse and underlie synaptic transmission and hence cognition. Here we consider the evidence that AP causes cognitive deficits through altering calcium homeostasis at the synapse, thus impairing synaptic transmission and LTP. Furthermore, this disruption appearr to occur without overt or extensive neuronal loss, as it is observed in transgenic mouse models of AD, but may contribute to the synaptic loss, which is an early event that correlates best with cognitive decline.

Hypoxic regulation of Ca2+ signalling in astrocytes and endothelial cells.

Acute hypoxia is well known to modulate plasmalemmal ion channels in specific tissue types, thereby modulating [Ca2+]i. Alternative mechanisms by which acute hypoxia could modulate [Ca2+]i are less well explored, particularly in non-excitable cells. Here, we describe experiments employing microfluorimetric recordings from Fura-2-loaded rat cortical astrocytes and human saphenous vein endothelial cells designed to explore any effects of hypoxia (pO2 20-30 mmHg) on [Ca2+]i. In both cell types, hypoxia evoked small rises of [Ca2+]i in the majority of cells during perfusion with a Ca(2+)-free solution, indicating hypoxia can release Ca2+ from an intracellular pool. Capacitative Ca2+ entry was observed when Ca2+ was subsequently restored to the extracellular solution. These effects were abolished by pre-treatment of cells with thapsigargin or prior application of inositol 1,4,5-trisphosphate (IP3)-generating agonists. Antioxidants fully prevented this effect of hypoxia in both cell types. Mitochondrial uncoupling significantly enhanced the effects of hypoxia in astrocytes, yet markedly suppressed the effects of hypoxia in endothelial cells. Our findings indicate that hypoxia can modulate [Ca2+]i in non-excitable cells; most importantly, it can evoke Ca2+ release from intracellular stores via a mechanism which involves reactive oxygen species. The involvement of mitochondria in this effect appears to be tissue specific.

Hypoxic regulation of Ca2+ signaling in cultured rat astrocytes.

Acute hypoxia modulates various cell processes, such as cell excitability, through the regulation of ion channel activity. Given the central role of Ca2+ signaling in the physiological functioning of astrocytes, we have investigated how acute hypoxia regulates such signaling, and compared results with those evoked by bradykinin (BK), an agonist whose ability to liberate Ca2+ from intracellular stores is well documented. In Ca2+-free perfusate, BK evoked rises of [Ca2+]i in all cells examined. Hypoxia produced smaller rises of [Ca2+]i in most cells, but always suppressed subsequent rises of [Ca2+]i induced by BK. Thapsigargin pre-treatment of cells prevented any rise of [Ca2+]i evoked by either BK or hypoxia. Restoration of Ca2+ to the perfusate following a period of acute hypoxia always evoked capacitative Ca2+ entry. During mitochondrial inhibition (due to exposure to carbonyl cyanide p-trifluromethoxyphenyl hydrazone (FCCP) and oligomycin), rises in [Ca2+]i (observed in Ca2+-free perfusate) evoked by hypoxia or by BK, were significantly enhanced, and hypoxia always evoked responses. Our data indicate that hypoxia triggers Ca2+ release from endoplasmic reticulum stores, efficiently buffered by mitochondria. Such liberation of Ca2+ is sufficient to trigger capacitative Ca2+ entry. These findings indicate that the local O2 level is a key determinant of astrocyte Ca2+ signaling, likely modulating Ca2+-dependent astrocyte functions in the central nervous system.

Chronic hypoxia potentiates capacitative Ca2+ entry in type-I cortical astrocytes.

Prolonged hypoxia exerts profound effects on cell function, and has been associated with increased production of amyloid beta peptides (A beta Ps) of Alzheimer's disease. Here, we have investigated the effects of chronic hypoxia (2.5% O2, 24 h) on capacitative Ca2+ entry (CCE) in primary cultures of rat type-I cortical astrocytes, and compared results with those obtained in astrocytes exposed to A beta Ps. Chronic hypoxia caused a marked enhancement of CCE that was observed after intracellular Ca2+ stores were depleted by bradykinin application or by exposure to thapsigargin (1 microM). Exposure of cells for 24 h to 1 microM A beta P(1-40) did not alter CCE. Enhancement of CCE was not attributable to cell hyperpolarization, as chronically hypoxic cells were significantly depolarized as compared with controls. Mitochondrial inhibition [by FCCP (10 microM) and oligomycin (2.5 microg/mL)] suppressed CCE in all three cell groups, but more importantly there were no significant differences in the magnitude of CCE in the three astrocyte groups under these conditions. Similarly, the antioxidants melatonin and Trolox abolished the enhancement of CCE in hypoxic cells. Our results indicate that chronic hypoxia augments CCE in cortical type-I astrocytes, a finding which is not mimicked by A beta P(1-40) and appears to be dependent on altered mitochondrial function.

Ca(2+) stores and capacitative Ca(2+) entry in human neuroblastoma (SH-SY5Y) cells expressing a familial Alzheimer's disease presenilin-1 mutation.

Presenilins are involved in the proteolytic production of Alzheimer's amyloid peptides, but are also known to regulate Ca(2+) homeostasis in various cells types. In the present study, we examined intracellular Ca(2+) stores coupled to muscarinic receptors and capacitative Ca(2+) entry (CCE) in the human neuroblastoma SH-SY5Y cell line, and how these were modulated by over-expression of either wild-type presenilin 1 (PS1wt) or a mutant form of presenilin 1 (PS1 deltaE9) which predisposes to early-onset Alzheimer's disease. Ca(2+) stores discharged by application of 100 microM muscarine (in Ca(2+)-free perfusate) in PS1wt and PS1 DeltaE9 cells were significantly larger than those in control cells, as determined using Fura-2 microfluorimetry. Subsequent CCE, observed in the absence of muscarine when Ca(2+) was re-admitted to the perfusate, was unaffected in PS1wt cells, but significantly suppressed in PS1 deltaE9 cells. However, when Ca(2+) stores were fully depleted with thapsigargin, CCE was similar in all three cell groups. Western blots confirmed increased levels of PS1 in the transfected cells, but also demonstrated that the proportion of intact PS1 in the PS1 deltaE9 cells was far greater than in the other two cell groups. This study represents the first report of modulation of both Ca(2+) stores and CCE in a human, neurone-derived cell line, and indicates a distinct effect of the PS1 mutation deltaE9 over wild-type PS1.

Chronic hypoxia remodels voltage-gated Ca2+ entry in a human airway chemoreceptor cell line.

Arterial and airway chemoreceptors respond to acute hypoxia by depolarizing, thereby activating voltage-gated Ca2+ channels and so permitting Ca2+ entry to trigger transmitter release. Following periods of prolonged hypoxia, these cells undergo a form of remodelling which involves altered expression of ion channels. Here, we use microspectrofluorimetric recordings of voltage-gated Ca2+ entry (activated by exposure of cells to 50 mM K+) to show that chronic hypoxia suppresses such Ca2+ entry in model airway chemoreceptor (H146) cells. Furthermore, Ca2+ entry via L-type channels is suppressed, whilst entry via N-type channels is greatly enhanced. The suppressed response, together with dramatic remodelling of routes available for voltage-gated Ca2+ entry, is likely to alter significantly the acute O2 sensing properties of these cells.

Prion protein fragment 106-126 potentiates catecholamine secretion from PC-12 cells.

The toxic actions of scrapie prion protein (PrP(sc)) are poorly understood. We investigated the ability of the toxic PrP(sc) fragment 106-126 to interfere with evoked catecholamine secretion from PC-12 cells. Prion protein fragment 106-126 (PrP106-126) caused a time- and concentration-dependent augmentation of exocytosis due to the emergence of a Ca(2+) influx pathway resistant to Cd(2+) but sensitive to other inorganic cations. In control cells, secretion was dependent on Ca(2+) influx through L- and N-type Ca(2+) channels, but after exposure to PrP106-126, secretion was unaffected by N-type channel blockade. Instead, selective L-type channel blockade was as effective as Cd(2+) in suppressing secretion. Patch-clamp recordings revealed no change in total Ca(2+) current density in PrP106-126-treated cells or in the contribution to total current of L-type channels, but a small Cd(2+)-resistant current was found only in PrP106-126-treated cells. Thus PrP106-126 augments secretion by inducing a Cd(2+)-resistant Ca(2+) influx pathway and alters coupling of native Ca(2+) channels to exocytosis. These effects are likely contributory factors in the toxic cellular actions of PrP(sc).

Effects of chronic hypoxia on Ca(2+) stores and capacitative Ca(2+) entry in human neuroblastoma (SH-SY5Y) cells.

Microfluorimetric measurements of intracellular calcium ion concentration [Ca(2+)](i) were employed to examine the effects of chronic hypoxia (2.5% O(2), 24 h) on Ca(2+) stores and capacitative Ca(2+) entry in human neuroblastoma (SH-SY5Y) cells. Activation of muscarinic receptors evoked rises in [Ca(2+)](i) which were enhanced in chronically hypoxic cells. Transient rises of [Ca(2+)](i) evoked in Ca(2+)-free solutions were greater and decayed more slowly following exposure to chronic hypoxia. In control cells, these transient rises of [Ca(2+)](i) were also enhanced and slowed by removal of external Na(+), whereas the same manoeuvre did not affect responses in chronically hypoxic cells. Capacitative Ca(2+) entry, observed when re-applying Ca(2+) following depletion of intracellular stores, was suppressed in chronically hypoxic cells. Western blots revealed that presenilin-1 levels were unaffected by chronic hypoxia. Exposure of cells to amyloid beta peptide (1-40) also increased transient [Ca(2+)](i) rises, but did not mimic any other effects of chronic hypoxia. Our results indicate that chronic hypoxia causes increased filling of intracellular Ca(2+) stores, suppressed expression or activity of Na(+)/Ca(2+) exchange and reduced capacitative Ca(2+) entry. These effects are not attributable to increased amyloid beta peptide or presenilin-1 levels, but are likely to be important in adaptive cellular remodelling in response to prolonged hypoxic or ischemic episodes.

Differential coupling of voltage-gated Ca(2+) channels to catecholamine secretion from separate PC12 cell batches.

Amperometric recordings were employed to investigate the coupling of Ca(2+) channels to catecholamine secretion in two batches of pheochromocytoma (PC12) cells. In 'new' (freshly obtained) cells (PC12n cells), secretion was dependent on Ca(2+) influx through L-type and N-type Ca(2+) channels. By contrast, in 'aged' cells (maintained in liquid nitrogen for 6-8 years; PC12a cells), secretion was mostly dependent on Ca(2+) influx through N-type channels. Patch clamp recordings revealed that L-type channels accounted for only ca. 26% of total whole-cell current in PC12a cells (determined by blockade caused by 2 microM nifedipine). In contrast, nifedipine suppressed currents by ca. 59% in PC12n cells. Thus important differences in fundamental physiological properties can be observed in PC12 cell batches even when obtained from the same source and maintained under identical conditions.

Melatonin has no effect on tolerance to uncompensable heat stress in man.

This study examined whether a 5 mg dose of melatonin induced a lower rectal temperature (Tre) response at rest in both a cool and hot environment while wearing normal military combat clothing, and then examined the influence of this response on tolerance to exercise in the heat while wearing protective clothing. Nine men performed four randomly ordered trials involving 2 h of rest at ambient temperatures of either 23 degrees C or 40 degrees C followed by exercise at an ambient temperature of 40 degrees C. The double-blind ingestion of placebo or melatonin occurred after 30 min of rest. The mean Tre during rest at 23 degrees C had decreased significantly from 36.8 (SD 0.1) degrees C to 36.7 (SD 0.2) degrees C at 90 min following the ingestion of the drug, whereas values during the placebo trial did not change. The lower Tre response during the melatonin trial remained during the first 50 min of exercise in the heat while wearing the protective clothing. Since the final mean Tre at the end of exercise also was significantly reduced for the melatonin [39.0 (SD 0.4) degrees C] compared with the placebo [mean 39.1 (SD 0.3) degrees C] trial, tolerance times approximated 95 min in both conditions. During rest at 40 degrees C, melatonin did not affect the mean Tre response which increased significantly during the last 90 min from 36.9 (SD 0.1) degrees C to 37.3 (SD 0.1) degrees C. This increase in Tre during the rest period prior to donning the protective clothing decreased tolerance time approximately 30 min compared with the trials that had involved rest at 23 degrees C. Total heat storage summated over the rest and exercise periods was not different among the trials at 15 kJ x kg(-1). It was concluded that the small decrease in Tre following the ingestion of 5 mg of melatonin at rest in a cool environment had no influence on subsequent tolerance during uncompensable heat stress.

Long-term results after omental patch repair in patients with perforated duodenal ulcers: a 5- to 10-year follow-up study.

Ninety-two patients with perforated duodenal ulcers were treated at two hospitals in St. John's. Five patients were managed conservatively, and the remainder received omental patch closure. All were prescribed histamine receptor antagonists. No patient had a definitive ulcer operation. Seventy-five patients were followed up for 5 to 10 years. Fourteen patients who had been taking ulcerogenic medications had a low recurrence rate (7%); the other 61 patients who had not been taking ulcerogenic medications had a high recurrence rate (77%). Use of an omental patch is effective treatment for perforated duodenal ulcers and provides long-term benefit for patients whose perforations are associated with ulcerogenic medications. Selected patients who have not been taking ulcerogenic medication are better treated by definitive ulcer surgery at the time of perforation.

Palliative surgery for adenocarcinoma of the gastroesophageal junction.

Adenocarcinomas of the gastroesophageal junction are usually incurable at the time the patient is first seen. The charts of 21 consecutive patients with this condition were reviewed. All had histologically documented transmural extension of tumour and lymph-node or distant metastases. A variety of surgical techniques were used to restore the ability to swallow. From the charts of these 21 cases, the authors analysed the success in restoring swallowing, the length of hospital stay, the surgical death rate, complications, survival and the ability to swallow normally at the time of death. Restoration of normal or near-normal swallowing appears to be a realistic goal. Selection of an appropriate surgical option, with resection of the primary tumour when technically feasible, allows this to be done with negligible mortality and acceptable morbidity.

Plant protein rehabilitation diets and iron supplementation of the protein-energy malnourished child.

Thirty-eight children admitted to the paediatric ward of Obafemi Awolowo University Teaching Hospital, Ile-Ife, were treated using a local therapeutic regimen for protein-energy malnutrition (PEM). Serum transferrin concentration was measured at weekly intervals to determine the response to and nutritional adequacy of the dietary component of this therapeutic regimen. At discharge, weight for height measurements in the children showed a significant (P less than 0.001) increase in growth although the expected weight for height was not attained. Serum transferrin concentration rose significantly from a mean basal level of 132.1 +/- 17.2 mg/dl to 222.0 +/- 24.6 mg/dl at the end of the first week of hospitalization and at discharge was 403.2 +/- 27.8 mg/dl. Although not statistically significant, mortality was highest amongst children for whom iron supplementation started early on admission. Earlier studies on the aetiology and treatment of anaemia in PEM have argued in favour of routine treatment with iron to prevent the development of anaemia during the recovery phase, but data from this study suggest that iron therapy should not be instituted during the first week of treatment.

Plasma somatomedin-C in Nigerian malnourished children fed a vegetable protein rehabilitation diet.

Plasma somatomedin-C (pSm-C) was measured by immunoassay in Nigerian malnourished children treated with a mainly vegetable diet. In oedematous children, the mean intake was 4.31 +/- 0.23 g protein and 611 +/- 46 kJ per kg body weight per day, and in marasmic children 5.22 +/- 0.62 g protein and 795 +/- 131 kJ/kg body weight/d. PSm-C concentration (U/ml) was measured at weekly intervals to determine the response to this rehabilitation diet. By our assay the value for 39 normal children (age range 6-36 months) was 0.315 +/- 0.035 U/ml. The average initial level of pSm-C in the malnourished children was 0.19 +/- 0.03 (n = 24). The values were higher (P less than 0.05) in the 7 marasmic children (0.26 +/- 0.1) than in the 11 with oedema (0.15 +/- 0.02). Eight days after admission pSm-C had risen to 0.20 +/- 0.02 (n = 24) and at discharge after approximately 19 d, pSm-C concentration was normal, 0.30 +/- 0.05. In oedematous malnutrition, pSm-C level at discharge was lower than in marasmus, 0.27 +/- 0.06 (n = 17) compared with 0.37 +/- 0.06 (n = 7) (P less than 0.05). Because the childrens' stay in hospital was short (average 19 d), they were far from attaining normal weight for height by the time of discharge. However, they had gained on average 0.9 kg and their clinical condition was satisfactory. It is concluded that the vegetable-based diet produced satisfactory recovery, at least in the initial stages. Increases in pSm-C compared well with those found in an earlier study with milk-based diets.

Nutrition support and malnutrition in Nigeria.

PIP: In 1983, a nutritional support team was formed at the University of Ife-Ife, Nigeria, that used high calorie enteral mixtures successfully for dietary management of protein energy malnutrition (PEM) in children. PEM has several causes. Poverty is often cited, but the incidence of mild to severe PEM in children under 5 is higher in the Ivory Coast, Nigeria, Egypt and Sudan with per capita gross national product (GNP) above $400 than in Sierra Leone, India, Uganda, and Kenya with GNP below this amount. The consumption of legumes and oil seeds ward off kwashiorkor and marasmus, but in countries with traditional food practices they are not consumed in adequate amounts. Beans, groundnuts, melon seeds, and soya beans are cheap and produced in African and Asian countries. In Nigeria the traditional weaning food is a thin gruel made from maize, sorghum, or millet. Milk, groundnut paste, or sugar is not added. Legumes and other oil seeds are forbidden for children because of deep-rooted cultural practices that favor tubers. Longer duration of breast feeding protects infants from kwashiorkor or marasmus, but the recent drastic change in the pattern with early introduction of artificial feeding has resulted in early appearance of kwashiorkor or gastroenteritis. Low literacy of mothers is another factor, and it inversely correlated with infant mortality. The increase in the level of female literacy and maternal education in less developed countries is a major requirement from governments if they are to combat harmful food taboos. Since Williams associated maize diets with kwashiorkor in 1933, research has show energy deficiency more perilous than protein insufficiency in the treatment and prevention of PEM in these countries.^ieng

Sodium and potassium intakes amongst free-living adult Nigerians.

Sixteen adult male and female Nigerians, aged between 18 and 48 years, participated in a 7-d study designed to estimate mean daily intakes of sodium and potassium and to determine their food sources. Food logs were kept during the 7-d study period. Analysis of data showed a mean daily intake of 8.1 g table salt with a range of 3.8 to 18.1 g which provided a mean daily intake of 3.2 g sodium. Mean sodium intake from foods alone was 0.85 g +/- 0.08 while sodium nutrient density was 2.3 g/1000 kcal. Potassium intake was 2.86 g +/- 0.2 or 1.46 g/1000 kcal. Mean sodium/potassium ratio of the diets was 1.5. Discretionary use of salt was 82 per cent of total intake with table salt supplying 78 per cent. Cereals and products provided the highest proportion of natural sodium while starchy staples supplied over 40 per cent of the potassium in the diet.

Fat contents of meals and bioavailability of griseofulvin in man.

The bioavailability of griseofulvin was followed in twelve healthy volunteers by measuring the urinary excretion of the major metabolite 6-demethylgriseolfulvin, after each volunteer had ingested one 500 mg griseofulvin tablet under (1) fasting conditions, (2) immediately after a typical low-fat and (3) high-fat Nigerian meals. An increase of about 70 and 120% absorption occurred with the ingestion of the low-fat and high-fat meals respectively compared to the fasting state (P less than 0.01). The maximum excretion rates of the free metabolite (Vmax.) were also significantly increased (P less than 0.01) following consumption of low and high fat meals. Our results thus suggest that the higher the fat content of the meals the higher the enhancement of the bioavailability of griseofulvin in man.