Expression of the vesicular GABA transporter within neuromedin S+ neurons sustains behavioral circadian rhythms.

The suprachiasmatic nucleus (SCN) of the hypothalamus is the site of a central circadian clock that orchestrates overt rhythms of physiology and behavior. Circadian timekeeping requires intercellular communication among SCN neurons, and multiple signaling pathways contribute to SCN network coupling. Gamma-aminobutyric acid (GABA) is produced by virtually all SCN neurons, and previous work demonstrates that this transmitter regulates coupling in the adult SCN but is not essential for the nucleus to sustain overt circadian rhythms. Here, we show that the deletion of the gene that codes for the GABA vesicular transporter Vgat from neuromedin-S (NMS)+ neurons-a subset of neurons critical for SCN function-causes arrhythmia of locomotor activity and sleep. Further, NMS-Vgat deletion impairs intrinsic clock gene rhythms in SCN explants cultured ex vivo. Although vasoactive intestinal polypeptide (VIP) is critical for SCN function, Vgat deletion from VIP-expressing neurons did not lead to circadian arrhythmia in locomotor activity rhythms. Likewise, adult SCN-specific deletion of Vgat led to mild impairment of behavioral rhythms. Our results suggest that while the removal of GABA release from the adult SCN does not affect the pacemaker's ability to sustain overt circadian rhythms, its removal from a critical subset of neurons within the SCN throughout development removes the nucleus ability to sustain circadian rhythms. Our findings support a model in which SCN GABA release is critical for the developmental establishment of intercellular network properties that define the SCN as a central pacemaker.

On the origin and evolution of the dual oscillator model underlying the photoperiodic clockwork in the suprachiasmatic nucleus.

Decades have now passed since Colin Pittendrigh first proposed a model of a circadian clock composed of two coupled oscillators, individually responsive to the rising and setting sun, as a flexible solution to the challenge of behavioral and physiological adaptation to the changing seasons. The elegance and predictive power of this postulation has stimulated laboratories around the world in searches to identify and localize such hypothesized evening and morning oscillators, or sets of oscillators, in insects, rodents, and humans, with experimental designs and approaches keeping pace over the years with technological advances in biology and neuroscience. Here, we recount the conceptual origin and highlight the subsequent evolution of this dual oscillator model for the circadian clock in the mammalian suprachiasmatic nucleus; and how, despite our increasingly sophisticated view of this multicellular pacemaker, Pittendrigh's binary conception has remained influential in our clock models and metaphors.

Developmental patterning of peptide transcription in the central circadian clock in both sexes.

Introduction: Neuropeptide signaling modulates the function of central clock neurons in the suprachiasmatic nucleus (SCN) during development and adulthood. Arginine vasopressin (AVP) and vasoactive intestinal peptide (VIP) are expressed early in SCN development, but the precise timing of transcriptional onset has been difficult to establish due to age-related changes in the rhythmic expression of each peptide. Methods: To provide insight into spatial patterning of peptide transcription during SCN development, we used a transgenic approach to define the onset of Avp and Vip transcription. Avp-Cre or Vip-Cre males were crossed to Ai9+/+ females, producing offspring in which the fluorescent protein tdTomato (tdT) is expressed at the onset of Avp or Vip transcription. Spatial patterning of Avp-tdT and Vip-tdT expression was examined at critical developmental time points spanning mid-embryonic age to adulthood in both sexes. Results: We find that Avp-tdT and Vip-tdT expression is initiated at different developmental time points in spatial subclusters of SCN neurons, with developmental patterning that differs by sex. Conclusions: These data suggest that SCN neurons can be distinguished into further subtypes based on the developmental patterning of neuropeptide expression, which may contribute to regional and/or sex differences in cellular function in adulthood.

Somatostatin regulates central clock function and circadian responses to light.

Daily and annual changes in light are processed by central clock circuits that control the timing of behavior and physiology. The suprachiasmatic nucleus (SCN) in the anterior hypothalamus processes daily photic inputs and encodes changes in day length (i.e., photoperiod), but the SCN circuits that regulate circadian and photoperiodic responses to light remain unclear. Somatostatin (SST) expression in the hypothalamus is modulated by photoperiod, but the role of SST in SCN responses to light has not been examined. Our results indicate that SST signaling regulates daily rhythms in behavior and SCN function in a manner influenced by sex. First, we use cell-fate mapping to provide evidence that SST in the SCN is regulated by light via de novo Sst activation. Next, we demonstrate that Sst  -/- mice display enhanced circadian responses to light, with increased behavioral plasticity to photoperiod, jetlag, and constant light conditions. Notably, lack of Sst  -/- eliminated sex differences in photic responses due to increased plasticity in males, suggesting that SST interacts with clock circuits that process light differently in each sex. Sst  -/- mice also displayed an increase in the number of retinorecipient neurons in the SCN core, which express a type of SST receptor capable of resetting the molecular clock. Last, we show that lack of SST signaling modulates central clock function by influencing SCN photoperiodic encoding, network after-effects, and intercellular synchrony in a sex-specific manner. Collectively, these results provide insight into peptide signaling mechanisms that regulate central clock function and its response to light.

Human genetic variant E756del in the ion channel PIEZO1 not associated with protection from severe malaria in a large Ghanaian study.

Recently, a common genetic variant E756del in the human gene PIEZO1 was associated with protection from severe malaria. Here, we performed a genetic association study of this gain-of-function variant in a large case-control study including 4149 children from the Ashanti Region in Ghana, West Africa. The statistical analysis did not indicate an association with protection from severe malaria and, thus, providing evidence against a strong protective effect of the PIEZO1 E756del variant on severe malaria susceptibility.

Sex differences in daily timekeeping and circadian clock circuits.

The circadian system regulates behavior and physiology in many ways important for health. Circadian rhythms are expressed by nearly every cell in the body, and this large system is coordinated by a central clock in the suprachiasmatic nucleus (SCN). Sex differences in daily rhythms are evident in humans and understanding how circadian function is modulated by biological sex is an important goal. This review highlights work examining effects of sex and gonadal hormones on daily rhythms, with a focus on behavior and SCN circuitry in animal models commonly used in pre-clinical studies. Many questions remain in this area of the field, which would benefit from further work investigating this topic.

Vasopressin Resets the Central Circadian Clock in a Manner Influenced by Sex and Vasoactive Intestinal Polypeptide Signaling.

BACKGROUND/AIMS: Circadian rhythms in behavior and physiology are programmed by the suprachiasmatic nucleus (SCN) of the hypothalamus. A subset of SCN neurons produce the neuropeptide arginine vasopressin (AVP), but it remains unclear whether AVP signaling influences the SCN clock directly. METHODS: Here, we test that AVP signaling acting through V1A and V1B receptors influences molecular rhythms in SCN neurons. V1 receptor agonists were applied ex vivo to PERIOD2::LUCIFERASE SCN slices, allowing for real-time monitoring of changes in molecular clock function. RESULTS: V1A/B agonists reset the phase of the SCN molecular clock in a time-dependent manner, with larger magnitude responses by the female SCN. Further, we found evidence that both Gαq and Gαs signaling pathways interact with V1A/B-induced SCN resetting, and that this response requires vasoactive intestinal polypeptide (VIP) signaling. CONCLUSIONS: Collectively, this work indicates that AVP signaling resets SCN molecular rhythms in conjunction with VIP signaling and in a manner influenced by sex. This highlights the utility of studying clock function in both sexes and suggests that signal integration in central clock circuits regulates emergent properties important for the control of daily rhythms in behavior and physiology.

Gastroenteritis Rehydration Of children with Severe Acute Malnutrition (GASTROSAM): A Phase II Randomised Controlled trial: Trial Protocol.

Background: Children hospitalised with severe acute malnutrition (SAM) are frequently complicated (>50%) by diarrhoea ( ≥3 watery stools/day) which is accompanied by poor outcomes. Rehydration guidelines for SAM are exceptionally conservative and controversial, based upon expert opinion. The guidelines only permit use of intravenous fluids for cases with advanced shock and exclusive use of low sodium intravenous and oral rehydration solutions (ORS) for fear of fluid and/or sodium overload. Children managed in accordance to these guidelines have a very high mortality. The proposed GASTROSAM trial is the first step in reappraising current recommendations. We hypothesize that liberal rehydration strategies for both intravenous and oral rehydration in SAM children with diarrhoea may reduce adverse outcomes. Methods An open Phase II trial, with a partial factorial design, enrolling Ugandan and Kenyan children aged 6 months to 12 years with SAM hospitalised with gastroenteritis (>3 loose stools/day) and signs of moderate and severe dehydration.  In Stratum A (severe dehydration) children will be randomised (1:1:2) to WHO plan C (100mls/kg Ringers Lactate (RL) with intravenous rehydration given over 3-6 hours according to age including boluses for shock), slow rehydration (100 mls/kg RL over 8 hours (no boluses)) or  WHO SAM rehydration regime (ORS only (boluses for shock (standard of care)).  Stratum B incorporates all children with moderate dehydration and severe dehydration post-intravenous rehydration and compares (1:1 ratio) standard WHO ORS given for non-SAM (experimental) versus WHO SAM-recommended low-sodium ReSoMal. The primary outcome for intravenous rehydration is urine output (mls/kg/hour at 8 hours post-randomisation), and for oral rehydration a change in sodium levels at 24 hours post-randomisation. This trial will also generate feasibility, safety and preliminary data on survival to 28 days. Discussion. If current rehydration strategies for non-malnourished children are safe in SAM this could prompt future evaluation in Phase III trials.

Incidence and predictors of hospital readmission in children presenting with severe anaemia in Uganda and Malawi: a secondary analysis of TRACT trial data.

BACKGROUND: Severe anaemia (haemoglobin < 6 g/dL) is a leading cause of recurrent hospitalisation in African children. We investigated predictors of readmission in children hospitalised with severe anaemia in the TRACT trial (ISRCTN84086586) in order to identify potential future interventions. METHODS: Secondary analyses of the trial examined 3894 children from Uganda and Malawi surviving a hospital episode of severe anaemia. Predictors of all-cause readmission within 180 days of discharge were identified using multivariable regression with death as a competing risk. Groups of children with similar characteristics were identified using hierarchical clustering. RESULTS: Of the 3894 survivors 682 (18%) were readmitted; 403 (10%) had ≥2 re-admissions over 180 days. Three main causes of readmission were identified: severe anaemia (n = 456), malaria (n = 252) and haemoglobinuria/dark urine syndrome (n = 165). Overall, factors increasing risk of readmission included HIV-infection (hazard ratio 2.48 (95% CI 1.63-3.78), p < 0.001); ≥2 hospital admissions in the preceding 12 months (1.44(1.19-1.74), p < 0.001); history of transfusion (1.48(1.13-1.93), p = 0.005); and missing ≥1 trial medication dose (proxy for care quality) (1.43 (1.21-1.69), p < 0.001). Children with uncomplicated severe anaemia (Hb 4-6 g/dL and no severity features), who never received a transfusion (per trial protocol) during the initial admission had a substantially lower risk of readmission (0.67(0.47-0.96), p = 0.04). Malaria (among children with no prior history of transfusion) (0.60(0.47-0.76), p < 0.001); younger-age (1.07 (1.03-1.10) per 1 year younger, p < 0.001) and known sickle cell disease (0.62(0.46-0.82), p = 0.001) also decreased risk of readmission. For anaemia re-admissions, gross splenomegaly and enlarged spleen increased risk by 1.73(1.23-2.44) and 1.46(1.18-1.82) respectively compared to no splenomegaly. Clustering identified four groups of children with readmission rates from 14 to 20%. The cluster with the highest readmission rate was characterised by very low haemoglobin (mean 3.6 g/dL). Sickle Cell Disease (SCD) predominated in two clusters associated with chronic repeated admissions or severe, acute presentations in largely undiagnosed SCD. The final cluster had high rates of malaria (78%), severity signs and very low platelet count, consistent with acute severe malaria. CONCLUSIONS: Younger age, HIV infection and history of previous hospital admissions predicted increased risk of readmission. However, no obvious clinical factors for intervention were identified. As missing medication doses was highly predictive, attention to care related factors may be important. TRIAL REGISTRATION: ISRCTN ISRCTN84086586 .

Collection of Mouse Brain Slices for Bioluminescence Imaging of Circadian Clock Networks.

Circadian rhythms in cellular function can be monitored in real time with bioluminescence imaging. In this approach, bioluminescence is produced by an enzymatic reaction, which can be used to report dynamic changes in gene or protein expression in living cells. Bioluminescence imaging in circadian experiments typically uses an ex vivo slice preparation, with the most commonly studied structure being the master clock in the suprachiasmatic nucleus (SCN) of the anterior hypothalamus. Here we describe procedures for dissecting and collecting SCN slices for bioluminescence imaging experiments.

Vasopressin regulates daily rhythms and circadian clock circuits in a manner influenced by sex.

Arginine vasopressin (AVP) is a neurohormone that alters cellular physiology through both endocrine and synaptic signaling. Circadian rhythms in AVP release and other biological processes are driven by the suprachiasmatic nucleus (SCN) of the anterior hypothalamus. Loss of vasopressin signaling alters circadian behavior, but the basis of these effects remains unclear. Here we investigate the role of AVP signaling in circadian timekeeping by analyzing behavior and SCN function in a novel AVP-deficient mouse model. Consistent with previous work, loss of AVP signaling increases water consumption and accelerates recovery to simulated jetlag. We expand on these results to show that loss of AVP increases period, imprecision and plasticity of behavioral rhythms under constant darkness. Interestingly, the effect of AVP deficiency on circadian period was influenced by sex, with loss of AVP lengthening period in females but not males. Examining SCN function directly with ex vivo bioluminescence imaging of clock protein expression, we demonstrate that loss of AVP signaling modulates the period, precision, and phase relationships of SCN neurons in both sexes. This pattern of results suggests that there are likely sex differences in downstream targets of the SCN. Collectively, this work indicates that AVP signaling modulates circadian circuits in a manner influenced by sex, which provides new insight into sexual dimorphisms in the regulation of daily rhythms.

Marking Time: Colorful New Insights into Master Clock Circuits.

A neural clock controls what we do each day, and understanding its circuitry is important for health. In this issue of Neuron, Shan et al. visualize molecular rhythms in subtypes of master clock neurons to test principles of cell identity and network wiring.

Reduced VIP Expression Affects Circadian Clock Function in VIP-IRES-CRE Mice (JAX 010908).

Circadian rhythms are programmed by the suprachiasmatic nucleus (SCN), which relies on neuropeptide signaling to maintain daily timekeeping. Vasoactive intestinal polypeptide (VIP) is critical for SCN function, but the precise role of VIP neurons in SCN circuits is not fully established. To interrogate their contribution to SCN circuits, VIP neurons can be manipulated specifically using the DNA-editing enzyme Cre recombinase. Although the Cre transgene is assumed to be inert by itself, we find that VIP expression is reduced in both heterozygous and homozygous adult VIP-IRES-Cre mice (JAX 010908). Compared with wild-type mice, homozygous VIP-Cre mice display faster reentrainment and shorter free-running period but do not become arrhythmic in constant darkness. Consistent with this phenotype, homozygous VIP-Cre mice display intact SCN PER2::LUC rhythms, albeit with altered period and network organization. We present evidence that the ability to sustain molecular rhythms in the VIP-Cre SCN is not due to residual VIP signaling; rather, arginine vasopressin signaling helps to sustain SCN function at both intracellular and intercellular levels in this model. This work establishes that the VIP-IRES-Cre transgene interferes with VIP expression but that loss of VIP can be mitigated by other neuropeptide signals to help sustain SCN function. Our findings have implications for studies employing this transgenic model and provide novel insight into neuropeptide signals that sustain daily timekeeping in the master clock.

Circuit development in the master clock network of mammals.

Daily rhythms are generated by the circadian timekeeping system, which is orchestrated by the master circadian clock in the suprachiasmatic nucleus (SCN) of mammals. Circadian timekeeping is endogenous and does not require exposure to external cues during development. Nevertheless, the circadian system is not fully formed at birth in many mammalian species and it is important to understand how SCN development can affect the function of the circadian system in adulthood. The purpose of the current review is to discuss the ontogeny of cellular and circuit function in the SCN, with a focus on work performed in model rodent species (i.e., mouse, rat, and hamster). Particular emphasis is placed on the spatial and temporal patterns of SCN development that may contribute to the function of the master clock during adulthood. Additional work aimed at decoding the mechanisms that guide circadian development is expected to provide a solid foundation upon which to better understand the sources and factors contributing to aberrant maturation of clock function.

Mortality risk over time after early fluid resuscitation in African children.

BACKGROUND: African children hospitalised with severe febrile illness have a high risk of mortality. The Fluid Expansion As Supportive Therapy (FEAST) trial (ISCRTN 69856593) demonstrated increased mortality risk associated with fluid boluses, but the temporal relationship to bolus therapy and underlying mechanism remains unclear. METHODS: In a post hoc retrospective analysis, flexible parametric models were used to compare change in mortality risk post-randomisation in children allocated to bolus therapy with 20-40 ml/kg 5% albumin or 0.9% saline over 1-2 h or no bolus (control, 4 ml/kg/hour maintenance), overall and for different terminal clinical events (cardiogenic, neurological, respiratory, or unknown/other). RESULTS: Two thousand ninety-seven and 1041 children were randomised to bolus vs no bolus, of whom 254 (12%) and 91 (9%) respectively died within 28 days. Median (IQR) bolus fluid in the bolus groups received by 4 h was 20 (20, 40) ml/kg and was the same at 8 h; total fluids received in bolus groups at 4 h and 8 h were 38 (28, 43) ml/kg and 40 (30, 50) ml/kg, respectively. Total fluid volumes received in the control group by 4 h and 8 h were median (IQR) 10 (6, 15) ml/kg and 10 (10, 26) ml/kg, respectively. Mortality risk was greatest 30 min post-randomisation in both groups, declining sharply to 4 h and then more slowly to 28 days. Maximum mortality risk was similar in bolus and no bolus groups; however, the risk declined more slowly in the bolus group, with significantly higher mortality risk compared to the no bolus group from 1.6 to 101 h (4 days) post-randomisation. The delay in decline in mortality risk in the bolus groups was most pronounced for cardiogenic modes of death. CONCLUSIONS: The increased risk from bolus therapy was not due to a mechanism occurring immediately after bolus administration. Excess mortality risk in the bolus group resulted from slower decrease in mortality risk over the ensuing 4 days. Thus, administration of modest bolus volumes appeared to prevent mortality risk declining at the same rate that it would have done without a bolus, rather than harm associated with bolus resulting from a concurrent increased risk of death peri-bolus administration. TRIAL REGISTRATION: ISRCTN69856593. Date of registration 15 December 2008.

Co-trimoxazole or multivitamin multimineral supplement for post-discharge outcomes after severe anaemia in African children: a randomised controlled trial.

BACKGROUND: Severe anaemia is a leading cause of paediatric admission to hospital in Africa; post-discharge outcomes remain poor, with high 6-month mortality (8%) and re-admission (17%). We aimed to investigate post-discharge interventions that might improve outcomes. METHODS: Within the two-stratum, open-label, multicentre, factorial randomised TRACT trial, children aged 2 months to 12 years with severe anaemia, defined as haemoglobin of less than 6 g/dL, at admission to hospital (three in Uganda, one in Malawi) were randomly assigned, using sequentially numbered envelopes linked to a second non-sequentially numbered set of allocations stratified by centre and severity, to enhanced nutritional supplementation with iron and folate-containing multivitamin multimineral supplements versus iron and folate alone at treatment doses (usual care), and to co-trimoxazole versus no co-trimoxazole. All interventions were administered orally and were given for 3 months after discharge from hospital. Separately reported randomisations investigated transfusion management. The primary outcome was 180-day mortality. All analyses were done in the intention-to-treat population; follow-up was 180 days. This trial is registered with the International Standard Randomised Controlled Trial registry, ISRCTN84086586, and follow-up is complete. FINDINGS: From Sept 17, 2014, to May 15, 2017, 3983 eligible children were randomly assigned to treatment, and followed up for 180 days. 164 (4%) were lost to follow-up. 1901 (95%) of 1997 assigned multivitamin multimineral supplement, 1911 (96%) of 1986 assigned iron and folate, and 1922 (96%) of 1994 assigned co-trimoxazole started treatment. By day 180, 166 (8%) children in the multivitamin multimineral supplement group versus 169 (9%) children in the iron and folate group had died (hazard ratio [HR] 0·97, 95% CI 0·79-1·21; p=0·81) and 172 (9%) who received co-trimoxazole versus 163 (8%) who did not receive co-trimoxazole had died (HR 1·07, 95% CI 0·86-1·32; p=0·56). We found no evidence of interactions between these randomisations or with transfusion randomisations (p>0·2). By day 180, 489 (24%) children in the multivitamin multimineral supplement group versus 509 (26%) children in the iron and folate group (HR 0·95, 95% CI 0·84-1·07; p=0·40), and 500 (25%) children in the co-trimoxazole group versus 498 (25%) children in the no co-trimoxazole group (1·01, 0·89-1·15; p=0·85) had had one or more serious adverse events. Most serious adverse events were re-admissions, occurring in 692 (17%) children (175 [4%] with at least two re-admissions). INTERPRETATION: Neither enhanced supplementation with multivitamin multimineral supplement versus iron and folate treatment or co-trimoxazole prophylaxis improved 6-month survival. High rates of hospital re-admission suggest that novel interventions are urgently required for severe anaemia, given the burden it places on overstretched health services in Africa. FUNDING: Medical Research Council and Department for International Development.

Transfusion Volume for Children with Severe Anemia in Africa.

BACKGROUND: Severe anemia (hemoglobin level, <6 g per deciliter) is a leading cause of hospital admission and death in children in sub-Saharan Africa. The World Health Organization recommends transfusion of 20 ml of whole-blood equivalent per kilogram of body weight for anemia, regardless of hemoglobin level. METHODS: In this factorial, open-label trial, we randomly assigned Ugandan and Malawian children 2 months to 12 years of age with a hemoglobin level of less than 6 g per deciliter and severity features (e.g., respiratory distress or reduced consciousness) to receive immediate blood transfusion with 20 ml per kilogram or 30 ml per kilogram. Three other randomized analyses investigated immediate as compared with no immediate transfusion, the administration of postdischarge micronutrients, and postdischarge prophylaxis with trimethoprim-sulfamethoxazole. The primary outcome was 28-day mortality. RESULTS: A total of 3196 eligible children (median age, 37 months; 2050 [64.1%] with malaria) were assigned to receive a transfusion of 30 ml per kilogram (1598 children) or 20 ml per kilogram (1598 children) and were followed for 180 days. A total of 1592 children (99.6%) in the higher-volume group and 1596 (99.9%) in the lower-volume group started transfusion (median, 1.2 hours after randomization). The mean (±SD) volume of total blood transfused per child was 475±385 ml and 353±348 ml, respectively; 197 children (12.3%) and 300 children (18.8%) in the respective groups received additional transfusions. Overall, 55 children (3.4%) in the higher-volume group and 72 (4.5%) in the lower-volume group died before 28 days (hazard ratio, 0.76; 95% confidence interval [CI], 0.54 to 1.08; P = 0.12 by log-rank test). This finding masked significant heterogeneity in 28-day mortality according to the presence or absence of fever (>37.5°C) at screening (P=0.001 after Sidak correction). Among the 1943 children (60.8%) without fever, mortality was lower with a transfusion volume of 30 ml per kilogram than with a volume of 20 ml per kilogram (hazard ratio, 0.43; 95% CI, 0.27 to 0.69). Among the 1253 children (39.2%) with fever, mortality was higher with 30 ml per kilogram than with 20 ml per kilogram (hazard ratio, 1.91; 95% CI, 1.04 to 3.49). There was no evidence of differences between the randomized groups in readmissions, serious adverse events, or hemoglobin recovery at 180 days. CONCLUSIONS: Overall mortality did not differ between the two transfusion strategies. (Funded by the Medical Research Council and Department for International Development, United Kingdom; TRACT Current Controlled Trials number, ISRCTN84086586.).

Immediate Transfusion in African Children with Uncomplicated Severe Anemia.

BACKGROUND: The World Health Organization recommends not performing transfusions in African children hospitalized for uncomplicated severe anemia (hemoglobin level of 4 to 6 g per deciliter and no signs of clinical severity). However, high mortality and readmission rates suggest that less restrictive transfusion strategies might improve outcomes. METHODS: In this factorial, open-label, randomized, controlled trial, we assigned Ugandan and Malawian children 2 months to 12 years of age with uncomplicated severe anemia to immediate transfusion with 20 ml or 30 ml of whole-blood equivalent per kilogram of body weight, as determined in a second simultaneous randomization, or no immediate transfusion (control group), in which transfusion with 20 ml of whole-blood equivalent per kilogram was triggered by new signs of clinical severity or a drop in hemoglobin to below 4 g per deciliter. The primary outcome was 28-day mortality. Three other randomizations investigated transfusion volume, postdischarge supplementation with micronutrients, and postdischarge prophylaxis with trimethoprim-sulfamethoxazole. RESULTS: A total of 1565 children (median age, 26 months) underwent randomization, with 778 assigned to the immediate-transfusion group and 787 to the control group; 984 children (62.9%) had malaria. The children were followed for 180 days, and 71 (4.5%) were lost to follow-up. During the primary hospitalization, transfusion was performed in all the children in the immediate-transfusion group and in 386 (49.0%) in the control group (median time to transfusion, 1.3 hours vs. 24.9 hours after randomization). The mean (±SD) total blood volume transfused per child was 314±228 ml in the immediate-transfusion group and 142±224 ml in the control group. Death had occurred by 28 days in 7 children (0.9%) in the immediate-transfusion group and in 13 (1.7%) in the control group (hazard ratio, 0.54; 95% confidence interval [CI], 0.22 to 1.36; P = 0.19) and by 180 days in 35 (4.5%) and 47 (6.0%), respectively (hazard ratio, 0.75; 95% CI, 0.48 to 1.15), without evidence of interaction with other randomizations (P>0.20) or evidence of between-group differences in readmissions, serious adverse events, or hemoglobin recovery at 180 days. The mean length of hospital stay was 0.9 days longer in the control group. CONCLUSIONS: There was no evidence of differences in clinical outcomes over 6 months between the children who received immediate transfusion and those who did not. The triggered-transfusion strategy in the control group resulted in lower blood use; however, the length of hospital stay was longer, and this strategy required clinical and hemoglobin monitoring. (Funded by the Medical Research Council and Department for International Development; TRACT Current Controlled Trials number, ISRCTN84086586.).

Gastroenteritis aggressive versus slow treatment for rehydration (GASTRO): a phase II rehydration trial for severe dehydration: WHO plan C versus slow rehydration.

BACKGROUND: World Health Organization rehydration management guidelines (plan C) for severe dehydration are widely practiced in resource-poor settings, but never formally evaluated in a trial. The Fluid Expansion as a Supportive Therapy trial raised concerns regarding the safety of bolus therapy for septic shock, warranting a formal evaluation of rehydration therapy for gastroenteritis. METHODS: A multi-centre open-label phase II randomised controlled trial evaluated two rehydration strategies in 122 Ugandan/Kenyan children aged 60 days to 12 years with severe dehydration secondary to gastroenteritis. We compared the safety and efficacy of standard rapid rehydration using Ringer's lactate (100 ml/kg over 3 h (6 h if < 1 year), incorporating 0.9% saline boluses for children with shock (plan C) versus slower rehydration: 100 ml/kg Ringer's lactate over 8 h (all ages) without boluses (slow: experimental). The primary outcome was the frequency of serious adverse events (SAE) within 48 h including cardiovascular, respiratory and neurological complications. Secondary outcomes included clinical, biochemical and physiological measures of response to treatment by intravenous rehydration. RESULTS: One hundred twenty-two eligible children (median (IQR) age 8 (6-12) months) were randomised to plan C (n = 61) or slow (n = 61), with two (2%) lost to follow-up at day 7). Following randomisation mean (SD) time to start intravenous rehydration started was 15 min (18) in both arms. Mean (SD) fluid received by 1 hour was greater in plan C (mean 20.2 ml/kg (12.2) and 33.1 ml/kg (17) for children < 1 year and >- 1 year respectively) versus 10.4 ml/kg (6.6) in slow arm. By 8 hours volume received were similar mean (SD) plan C: 96.3 ml/kg (15.6) and 97.8 ml/kg (10.0) for children < 1 and ≥ 1 year respectively vs 93.2 ml/kg (12.2) in slow arm. By 48-h, three (5%) plan C vs two (3%) slow had an SAE (risk ratio 0.67, 95% CI 0.12-3.85, p = 0.65). There was no difference in time to the correction of dehydration (p = 0.9) or time to discharge (p = 0.8) between groups. Atrial natriuretic peptide levels rose substantially by 8 hours in both arms, which persisted to day 7. Day 7 weights suggested only 33 (29%) could be retrospectively classified as severely dehydration (≥ 10% weight loss). CONCLUSION: Slower rehydration over 8 hours appears to be safe, easier to implement than plan C. Future large trials with mortality as the primary endpoint are warranted. TRIAL REGISTRATION: ISRCTN67518332 . Date applied 31 August 2016.