Tunneling Spectroscopy at Megabar Pressures: Determination of the Superconducting Gap in Sulfur.

The recent discovery of high-temperature, high-pressure superconductors, such as hydrides and nickelates, has opened exciting avenues in studying high-temperature superconductivity. The primary superconducting properties of these materials are well characterized by measuring various electrical and magnetic properties, despite the challenges posed by the high-pressure environment. Experimental microscopic insight into the pairing mechanism of these superconductors is even more challenging, due to the lack of direct probes of the superconducting gap structures at high pressure conditions. Here, we have developed a planar tunnel junction technique for diamond anvil cells and present ground-breaking tunneling spectroscopy measurements at megabar pressures. We determined the superconducting gap of elemental sulfur at 160 GPa, a key constituent of the high-temperature superconductor H_{3}S. High quality tunneling spectra indicate that ß-Po phase sulfur is a type II superconductor with a single s-wave gap with a gap value 2Δ(0)=5.6 meV. This technique is compatible with superconducting compounds synthesized in diamond anvil cells and provides insight into the pairing mechanism in novel superconductors under high-pressure conditions.

Ontogenetic allometry and architectural properties of the paravertebral and hindlimb musculature in Eastern cottontail rabbits (Sylvilagus floridanus): functional implications for developmental changes in locomotor performance.

Due to small body size, an immature musculoskeletal system, and other growth-related limits on performance, juvenile mammals frequently experience a greater risk of predation than their adult counterparts. As a result, behaviorally precocious juveniles are hypothesized to exhibit musculoskeletal advantages that permit them to accelerate rapidly and evade predation. This hypothesis was tested through detailed quantitative evaluation of muscle growth in wild Eastern cottontail rabbits (Sylvilagus floridanus). Cottontail rabbits experience high rates of mortality during the first year of life, suggesting that selection might act to improve performance in growing juveniles. Therefore, it was predicted that muscle properties associated with force and power capacity should be enhanced in juvenile rabbits to facilitate enhanced locomotor performance. We quantified muscle architecture from 24 paravertebral and hindlimb muscles across ontogeny in a sample of n = 29 rabbits and evaluated the body mass scaling of muscle mass (MM), physiological cross-sectional area (PCSA), isometric force (Fmax ), and instantaneous power (Pinst ), along with several dimensionless architectural indices. In contrast to our hypothesis, MM and PCSA for most muscles change with positive allometry during growth by scaling at Mb1.3 and Mb1.1 , respectively, whereas Fmax and Pinst generally scale indistinguishably from isometry, as do the architectural indices tested. However, scaling patterns indicate that the digital flexors and ankle extensors of juvenile S. floridanus have greater capacities for force and power, respectively, than those in adults, suggesting these muscle properties may be a part of several compensatory features that promote enhanced acceleration performance in young rabbits. Overall, our study implies that body size constraints place larger, more mature rabbits at a disadvantage during acceleration, and that adults must develop hypertrophied muscles in order to maintain mechanical similarity in force and power capacities across development. These findings challenge the accepted understanding that juvenile animals are at a performance detriment relative to adults. Instead, for prey-predator interactions necessitating short intervals of high force and power generation relative to body mass, as demonstrated by rapid acceleration of cottontail rabbits fleeing predators, it may be the adults that struggle to keep pace with juveniles.

Toxicokinetics and toxicity of atorvastatin in dogs.

HMG-CoA reductase inhibitors (e.g., statins) are an important clinical option to lower cholesterol and treat co-morbidities. Atorvastatin is the most prescribed statin and has obtained generic status. We recently had a clinical development program evaluating a combination of atorvastatin with a GPR119 agonist as a treatment for dyslipidemia, where toxicological evaluations in dogs were completed. There were several challenges related to selecting doses for atorvastatin, including understanding the dose-exposure relationship from different drug forms used by the innovator in their general toxicology studies, bioanalytical assays that did not separate and quantify parent from metabolites, and high variability in the systemic exposures following oral dosing. The studies in this report characterized the toxicokinetics and toxicity of atorvastatin in the dog for up to 13-weeks. Overall, there were no notable differences in the toxicokinetics of atorvastatin or the two active hydroxylated metabolites between the sexes at Week 13. However, systemic exposures were markedly lower at Week 13 compared to that observed at Week 4, suggesting induction of metabolism or reduced absorption from the gastrointestinal tract following oral dosing. Changes in laboratory chemistries included increased liver enzyme levels and lower cholesterol levels. Histopathologic evaluation revealed multifocal minimal to slight hemorrhages in the submucosa of the gallbladder; all findings were reversible. The information from these studies along with the existing clinical experience with atorvastatin can be used to design robust toxicology studies in dogs and reduce animal use.

Audit and review for evidence-based red cell wastage reduction measures.

Stocks of red blood cells (RBC) are held to ideally match supply and demand; hold too great a stock and unnecessary wastage occurs; too low a stock results in delay or lack of blood for the patient. Blood is a precious resource and its supply needs to be managed effectively. The aim was to identify how RBC units are wasted and propose laboratory-based reduction measures that would not compromise the clinical requirements of the patient. Wastage of RBC was investigated using a 'dashboard' query of a laboratory information management system. By employing service improvement tools, proposals were made to reduce unnecessary RBC waste while ensuring an adequate supply to the patient. The efficacy of those proposals was examined using the same dashboard to compare similar periods before and after their introduction. The reduction in RBC wastage for all groups during an eight month period (December to July) was from 6.4% (5.3% non-AB or B RhD-positive) pre-implementation to 4.4% (2.5% non-AB/B RhD-positive) post-implementation. Group O RhD-negative wastage reduced from 10.4% to 4.4% after introduction of waste-saving proposals. However, there was an increase in staff time required to introduce the changes and in associated Group and Screen testing (3.4 to 3.8 per unit issued). RBC wastage was significantly reduced (P<0.0001) by 32.8% (52%, non-AB/B RhD-positive), saving approximately 225 RBC units per annum. Financially, increased associated costs did not negate the savings made by the measures introduced.

Cdk12 maintains the integrity of adult axons by suppressing actin remodeling.

The role of cyclin-dependent kinases (CDKs) that are ubiquitously expressed in the adult nervous system remains unclear. Cdk12 is enriched in terminally differentiated neurons where its conical role in the cell cycle progression is redundant. We find that in adult neurons Cdk12 acts a negative regulator of actin formation, mitochondrial dynamics and neuronal physiology. Cdk12 maintains the size of the axon at sites proximal to the cell body through the transcription of homeostatic enzymes in the 1-carbon by folate pathway which utilize the amino acid homocysteine. Loss of Cdk12 leads to elevated homocysteine and in turn leads to uncontrolled F-actin formation and axonal swelling. Actin remodeling further induces Drp1-dependent fission of mitochondria and the breakdown of axon-soma filtration barrier allowing soma restricted cargos to enter the axon. We demonstrate that Cdk12 is also an essential gene for long-term neuronal survival and loss of this gene causes age-dependent neurodegeneration. Hyperhomocysteinemia, actin changes, and mitochondrial fragmentation are associated with several neurodegenerative conditions such as Alzheimer's disease and we provide a candidate molecular pathway to link together such pathological events.

Higher education does not protect against firework-related injuries: a review of the economic burden and the risk factors of firework-related injuries in the capital of Iran.

OBJECTIVE: To examine the incidence and risk factors of firework-related injuries during the Last Wednesday Eve Festival in Tehran, Iran, with a focus on the association of socio-economic status and educational level with the use of fireworks and the incidence of firework-related injury. STUDY DESIGN: Cross-sectional household survey. METHODS: Using a random cluster sampling approach, a household survey was conducted in Greater Tehran in April 2008. During a structured interview with an adult member of the household, questions were asked about the use of fireworks and any firework-related injuries sustained by household members during the preceding festival. Data were gathered on expenditure on fireworks, medical treatment of firework-related injuries, length of hospital stay for the treatment of these injuries, and damage to personal property by fireworks. RESULTS: The survey included 2456 households in Greater Tehran. At least one member of 18% of these households had used fireworks during the Last Wednesday Eve Festival in 2008. The overall incidence of firework-related injuries was 100 per 100,000 population (95% confidence interval 37-163). The use of fireworks was less common among parents and more common among male children. Individuals who used fireworks were younger than non-users. Younger age and use of fireworks were associated with firework-related injuries (P < 0.05). The mean household expenditure on fireworks was US$1.62. Among the households that had bought fireworks, the mean expenditure was US$9.40 (standard deviation US$16.34). Thirty-two households (1.3%) reported damage to personal property due to fireworks during the festival costing US$3.30-167.20. The regional price of housing in the study area was correlated with the educational level of the head of the household. Higher educational level of the head of the household was associated with participation in firework activities by household members, expenditure on fireworks, and the amount of financial loss due to fireworks (all P < 0.05). CONCLUSION: Fireworks are associated with serious injuries, and impose a non-trivial financial burden on families. While personal use of fireworks was an independent risk factor for firework-related injuries, higher socio-economic status of the household and higher educational level of the head of the household were not protective factors.

Context-driven changes in L-DOPA-induced behaviours in the 6-OHDA lesioned rat.

Both contralateral rotational behaviour and dyskinetic abnormal involuntary movements (AIMs) are induced by the administration of l-DOPA in the unilateral 6-OHDA lesioned rat model of Parkinson's disease. Since rotational responses can be conditioned to environmental cues we have investigated the extent to which drug-induced AIMS may also be conditioned by exteroceptive cues and experience. In Experiment I, 6-OHDA lesioned rats received repeated daily injections of l-DOPA either in their home cage (control) or in association with a brief (20 mins) exposure to the rotometers (paired). To assess conditioning, all animals then received two tests in the rotometer bowls. Following injection of saline the paired group both rotated more contralaterally and displayed manifest AIMs, neither of which were exhibited by the control rats. Moreover, following injection of l-DOPA, the paired group showed a trend for increased AIMs compared to controls. Two further studies provided longer exposure to the conditioning environments in counterbalanced designs. Although, using these parameters, re-exposure in the presence of saline did not induce context-dependent AIMs, a strong context-specific component of the sensitised response to l-DOPA was seen; chronic administration of drug produced a significantly stronger behavioural response in animals paired with a particular environment for drug administration than controls. This data suggests that part of the sensitisation of behavioural responding to l-DOPA administration is not solely a pharmacological phenomenon, but is also conditioned to the environmental context in which the drug is administered. This has clear implications for the clinical observation and experimental measurement of drug-induced dyskinesia in Parkinson's disease patients and animal models.

Benzodiazepine exposures among women of reproductive age in the US, 2004-2018.

Benzodiazepines, often used to treat anxiety, insomnia, and other conditions, are prescribed more frequently to women than men, and emergency department visits and overdose deaths involving benzodiazepines have increased significantly among women in recent years. This study describes characteristics and trends associated with benzodiazepine exposures among women of reproductive age (15-49 years old) that were reported to United States poison control centers from 2004 through 2018. The National Poison Data System recorded 258,370 first-ranked benzodiazepine exposures among women 15-49 years old during the study period. More than one-half (56.9%) of exposures involved a single-substance and one-third (34.0%) occurred among women 20-29 years old. The majority were categorized as "intentional, suspected suicide" (73.2%) or "intentional" (12.9%). Exposures frequently resulted in admission to a psychiatric facility (20.6%), critical care unit (18.1%), or non-critical care unit (9.3%). Twenty percent of cases resulted in a serious medical outcome, including 205 deaths. The substantial percentage of benzodiazepine exposures among women of reproductive age that were intentional and associated with suicide attempts or suicide deaths indicate that increased prevention efforts are needed to address this issue.

Injury-related childhood mortality in migrant households in a southern city of China.

BACKGROUND: Sporadic evidence from China suggests that migrant children are at higher risk of injury-related mortality than local indigenous children. METHODS: Child deaths from 2004 to 2008 were provided by the Shenzhen Women and Child Health Surveillance System. Population data for children 1-4 years old were obtained from the Shenzhen Bureau of Statistics and number of live births was obtained from birth registration records. All-cause and injury-related childhood mortality rates and death causes were calculated and compared. RESULTS: A total of 3774 deaths were identified. All-cause mortality rates per 10,000 dropped significantly from 66.28 (95% CI 60.50 to 72.06) in infants (<1 year old) and 7.40 (95% CI 6.16 to 8.64) in early childhood (1-4 years old) in 2004 to 40.42 (95% CI 37.31 to 43.53) and 3.97 (95% CI 3.36 to 4.58) in 2008. However, injury-related mortality rates did not change significantly from 2.36 (95% CI 1.27 to 3.45) in infants and 2.97 (95% CI 2.19 to 3.76) in early childhood in 2004 to 2.00 (95% CI 1.31 to 2.69) and 2.00 (95% CI 1.56 to 2.43) in 2008. Injury-related mortality rates were significantly higher among migrant children (p<0.05). Drowning and traffic crashes were the top two causes of early childhood injury deaths; suffocation was the leading cause of infant injury deaths. CONCLUSION: Migrant children were at significantly higher risk of injury-related mortality than local indigenous children. Injury prevention in Shenzhen should target drowning and traffic safety among young children and suffocation among infants as top priorities.

The tandem repeat domain in the Listeria monocytogenes ActA protein controls the rate of actin-based motility, the percentage of moving bacteria, and the localization of vasodilator-stimulated phosphoprotein and profilin.

The ActA protein is responsible for the actin-based movement of Listeria monocytogenes in the cytosol of eukaryotic cells. Analysis of mutants in which we varied the number of proline-rich repeats (PRR; consensus sequence DFPPPPTDEEL) revealed a linear relationship between the number of PRRs and the rate of movement, with each repeat contributing approximately 2-3 microns/min. Mutants lacking all functional PRRs (generated by deletion or point mutation) moved at rates 30% of wild-type. Indirect immunofluorescence indicated that the PRRs were directly responsible for binding of vasodilator-stimulated phosphoprotein (VASP) and for the localization of profilin at the bacterial surface. The long repeats, which are interdigitated between the PRRs, increased the frequency with which actin-based motility occurred by a mechanism independent of the PRRs, VASP, and profilin. Lastly, a mutant which expressed low levels of ActA exhibited a phenotype indicative of a threshold; there was a very low percentage of moving bacteria, but when movement did occur, it was at wild-type rates. These results indicate that the ActA protein directs at least three separable events: (1) initiation of actin polymerization that is independent of the repeat region; (2) initiation of movement dependent on the long repeats and the amount of ActA; and (3) movement rate dependent on the PRRs.

Synergistic signals in the mechanism of antigen-induced exocytosis in 2H3 cells: evidence for an unidentified signal required for histamine release.

The aim of this study was to determine whether the increase in cytosolic free Ca2+ concentration ([Ca2+]i) in response to antigen (aggregated ovalbumin) on IgE-primed 2H3 cells was sufficient to account for exocytosis. When the [Ca2+]i responses to antigen and the Ca2+ ionophore A23187 were compared, A23187 was much less effective at releasing histamine at equivalent [Ca2+]i increases, and little or no stimulated histamine release occurred with A23187 concentrations that matched the [Ca2+]i response to antigen concentrations that stimulated maximal histamine release. The [Ca2+]i response to antigen is not, therefore, sufficient to account for exocytosis, although extracellular Ca2+ is necessary to initiate both the [Ca2+]i response and histamine release: the antigen must generate an additional, unidentified, signal that is required for exocytosis. To determine whether this signal was the activation of protein kinase C, the effects of the phorbol ester 12-0-tetradecanoyl phorbol 13-acetate (TPA) on the responses to antigen were examined. TPA blocked the antigen-induced [Ca2+]i response and the release of inositol phosphates but had little effect on histamine release and did not stimulate exocytosis by itself. The unidentified signal from the antigen is therefore distinct from the activation of protein kinase C and is generated independently of the [Ca2+]i response or the release of inositol phosphates. Taken together with other data that imply that there is very little activation of protein kinase C by antigen when the rate of histamine release is maximal, it is concluded that the normal exocytotic response to antigen requires the synergistic action of the [Ca2+]i signal together with an unidentified signal that is not mediated by protein kinase C.

Unintentional pediatric submersion-injury-related hospitalizations in the United States, 2003.

The objective of this study was to examine the demographic characteristics and hospital resource utilization of submersion-injury-related hospitalizations among persons < or =20 years of age in the USA in 2003. All 1475 pediatric submersion-injury-related hospital discharges in the Kids' Inpatient Database were identified by ICD-9-CM diagnosis code or external cause of injury code. These cases represent an estimated 2490 pediatric submersion-injury-related hospitalizations nationwide. Inpatient costs for these estimated hospitalizations were approximately $10 million. The overall pediatric submersion-injury-related rate of hospitalization was 3.0 per 100,000 persons. Children aged 0-4 years had the highest rate of hospitalization (7.7 per 100,000 persons). Children with permanent submersion-injury-related morbidity accounted for 5.8% of hospital admissions and 37.3% of hospital costs in our study, and children with submersion-injury-related in-hospital death accounted for 11.6% of hospital admissions and 20.0% of hospital costs in our study. Prevention of submersion injury using focused, proven strategies deserves increased attention.

Personality disorders and nonfatal unintentional injuries among US adults.

OBJECTIVE: To investigate the association between personality disorders and nonfatal unintentional injuries in a representative sample of US adults. METHODS: Data on self-reported nonfatal unintentional injuries during the 12 months before the interview were obtained from the National Epidemiologic Survey on Alcohol and Related Conditions (NESARC) were analyzed; 43,093 adults > or = 18 years participated in the NESARC wave I survey in 2001-02. Personality disorders were determined using the NIAAA Alcohol Use Disorders and Associated Disabilities Interview Schedule-DSM-IV. RESULTS: Individuals with at least one personality disorder had a significantly higher 12-month incidence of injuries than people without any personality disorder (p<0.001). After accounting for sociodemographic characteristics or other mental disorders, OR was 1.54 (95% CI 1.39 to 1.71) for individuals with one personality disorder and 1.80 (95% CI 1.58 to 2.05) for individuals with two or more personality disorders compared with people with no personality disorder. CONCLUSION: Personality disorders were associated with a significantly increased risk of unintentional injuries. This information has important implications for the treatment of patients with these disorders.

Splanchnic insulin metabolism in obesity. Influence of body fat distribution.

The effects of obesity and body fat distribution on splanchnic insulin metabolism and the relationship to peripheral insulin sensitivity were assessed in 6 nonobese and 16 obese premenopausal women. When compared with the nonobese women, obese women had significantly greater prehepatic production and portal vein levels of insulin both basally and following glucose stimulation. This increase correlated with the degree of adiposity but not with waist-to-hip girth ratio (WHR). WHR, however, correlated inversely with the hepatic extraction fraction and directly with the posthepatic delivery of insulin. The latter correlated with the degree of peripheral insulinemia. The decline in hepatic insulin extraction with increasing WHR also correlated with the accompanying diminution in peripheral insulin sensitivity. Increasing adiposity is thus associated with insulin hypersecretion. The pronounced hyperinsulinemia of upper body fat localization, however, is due to an additional defect in hepatic insulin extraction. This defect is closely allied with the decline in peripheral insulin sensitivity.

JAK/STAT pathway inhibition overcomes IL7-induced glucocorticoid resistance in a subset of human T-cell acute lymphoblastic leukemias.

While outcomes for children with T-cell acute lymphoblastic leukemia (T-ALL) have improved dramatically, survival rates for patients with relapsed/refractory disease remain dismal. Prior studies indicate that glucocorticoid (GC) resistance is more common than resistance to other chemotherapies at relapse. In addition, failure to clear peripheral blasts during a prednisone prophase correlates with an elevated risk of relapse in newly diagnosed patients. Here we show that intrinsic GC resistance is present at diagnosis in early thymic precursor (ETP) T-ALLs as well as in a subset of non-ETP T-ALLs. GC-resistant non-ETP T-ALLs are characterized by strong induction of JAK/STAT signaling in response to interleukin-7 (IL7) stimulation. Removing IL7 or inhibiting JAK/STAT signaling sensitizes these T-ALLs, and a subset of ETP T-ALLs, to GCs. The combination of the GC dexamethasone and the JAK1/2 inhibitor ruxolitinib altered the balance between pro- and anti-apoptotic factors in samples with IL7-dependent GC resistance, but not in samples with IL7-independent GC resistance. Together, these data suggest that the addition of ruxolitinib or other inhibitors of IL7 receptor/JAK/STAT signaling may enhance the efficacy of GCs in a biologically defined subset of T-ALL.

Chapter Eight - Ubiquitin-Mediated Regulation of Cellular Responses to Vascular Endothelial Growth Factors.

Vascular endothelial growth factors (VEGFs) bind receptor tyrosine kinases (VEGFRs) to regulate vascular and lymphatic development and homeostasis. Such interactions are also implicated in pathological conditions ranging from cancer to heart disease. Increasingly, it is evident that ubiquitination plays a central role in regulating VEGFR function and the cellular response to VEGFs. E1, E2, and E3 ubiquitination enzymes deliver ubiquitin-specific modifications to protein substrates but there is much debate on the exact enzymes involved. The deubiquitinase (DUB) enzymes remove such modifications and are attracting increasing interest as potential therapeutic targets in a host of different disease states. Understanding how these enzyme families regulate VEGFR function in different cells and tissues is a major challenge. An understanding of the fundamental mechanisms underlying such biochemical regulation is needed for providing new therapeutics that target diseases such as cancer and heart disease.

Fibroblast Biomarkers of Sporadic Parkinson's Disease and LRRK2 Kinase Inhibition.

It has been uncertain whether specific disease-relevant biomarker phenotypes can be found using sporadic Parkinson's disease (PD) patient-derived samples, as it has been proposed that there may be a plethora of underlying causes and pathological mechanisms. Fibroblasts derived from familial PD patients harboring leucine-rich repeat kinase 2 (LRRK2), PTEN-induced putative kinase 1 (PINK1), and Parkin mutations show clear disease-relevant mitochondrial phenotypes, which are exacerbated under conditions of pharmacological stress. We utilized fibroblasts derived from non-familial sporadic PD patients (without LRRK2 mutations) or LRRK2 mutation carriers to directly compare the cellular phenotypes during and after mitochondrial stress. We then determined the effects of pharmacological LRRK2 kinase inhibition using LRRK2-in-1. We found that there were two distinct populations of sporadic PD patient-derived fibroblast lines. One group of sporadic PD lines was highly susceptible to valinomycin-induced mitochondrial depolarization, emulating the mutant LRRK2 phenotype. These lines showed elevated mitochondrial superoxide/ nitric oxide levels, displayed increased mitochondrial and lysosome co-localization, and an increased rate of mitochondrial collapse, which corresponded with changes in mitochondrial fission and fusion proteins. The application of LRRK2-in-1 reversed decreased levels of mitochondrial and lysosome co-localization and partially restored mitochondrial network associated proteins and the mitochondrial membrane potential in the fibroblasts. This study identifies novel mitochondrial biomarkers in sporadic PD patient-derived fibroblast lines, which could be used as preclinical tools in which to test novel and known neuroprotective compounds.

How the Listeria monocytogenes ActA protein converts actin polymerization into a motile force.

The ActA protein is an essential determinant of pathogenicity that is responsible for the actin-based motility of Listeria monocytogenes in mammalian cells and cell-free extracts. ActA appears to control at least four functions that collectively lead to actin-based motility: (1) initiation of actin polymerization, (2) polarization of ActA function, (3) transformation of actin polymerization into a motile force and (4) acceleration of movement mediated by the host protein profilin.

The effect of bilayer thickness on the activity of (Na+ + K+)-ATPase.

The activities of purified (Na+ + K+)-ATPase supported by a series of phosphatidylcholines with monounsaturated (cis-9) fatty acyl chains (di(n : 1) phosphatidylcholine) varying in length from n = 12 to n = 23 were determined by the lipid titration technique. The ATPase activity at 20 degrees C decreased from 2.9 to 0.1 mumol/min per mg protein as n was decreased from 16 to 12 and decreased from 2.9 to 1.0 mumol/min per mg protein as n was increased from 20 to 23. In further experiments, the di(n : 1) phosphatidylcholine-ATPase complexes were treated with increasing proportions of n-decane, which has been shown previously to increase the thickness of black lipid membranes. n-Decane caused a large increase (greater than 20-fold) in activity of the short-chain complexes (n = 12,13); for n = 14--18, the ATPase activity first increased and subsequently decreased as the proportion of decane was increased, and for n = 20 or 23 decane caused a progressive decrease in activity with increasing concentration. These effects confirm qualitatively that a major factor determining the activity in each bilayer is its thickness. This behaviour closely parallels that of the (Ca2+ + Mg2+)-ATPase of sarcoplasmic reticulum [1] and suggests that a major class of trans-membrane transport proteins may have a similar dependence on bilayer thickness.

Changes in free-calcium levels and pH in synaptosomes during transmitter release.

The free cytoplasmic Ca2+ concentration [( Ca2+]i) in rat brain synaptosomes estimated by the indicator quin 2 is 104 +/- 8 nM (S.D.) in artificial cerebrospinal fluid (1.2 mM Ca2+), but decreases at lower Ca2+ concentrations in the medium. The presence of quin 2 in the synaptosomes does not affect either the spontaneous release of transmitter (gamma-aminobutyric acid) or the release induced by K+ depolarisation. In quin 2-loaded synaptosomes, depolarisation by K+ causes an abrupt increase in [Ca]i (less than 2-fold) that is approximately proportional to the extent of depolarisation, whereas depolarisation by veratrine alkaloids produces a slow rise in [Ca]i. The increase in [Ca]i produced by K+ depolarisation does not occur in the absence of Ca2+ in the medium. The data are consistent with a direct correlation between [Cai] and transmitter release in functional synaptosomes. The pH in synaptosomes estimated by the indicator quene 1 is 7.04 +/- 0.07 and is stable in media containing 5 mM bicarbonate. The pH in synaptosomes was decreased by protoveratrine but not by K+ depolarisation.