Defining protein kinase/phosphatase isoenzymic regulation of mGlu5 receptor-stimulated phospholipase C and Ca²âº responses in astrocytes.

BACKGROUND AND PURPOSE: Cyclical phosphorylation and dephosphorylation of a key residue within the C-terminal domain of the activated type 5 metabotropic glutamate (mGlu5) receptor is believed to cause the synchronous, oscillatory changes in inositol 1,4,5-trisphosphate and Ca²âº levels observed in a variety of cell types. Here, we have attempted to better define the kinase and phosphatase enzymes involved in this modulation. EXPERIMENTAL APPROACH: Ca²âº and [³H]inositol phosphate ([³H]IP(x) ) measurements in astrocyte preparations have been used to evaluate the effects of pharmacological inhibition of protein kinase C (PKC) and protein phosphatase activities and small interfering RNA-mediated specific PKC isoenzymic knock-down on mGlu5 receptor signalling. KEY RESULTS: Ca²âº oscillation frequency or [³H]IP(x) accumulation in astrocytes stimulated by mGlu5 receptors, was concentration-dependently decreased by protein phosphatase-1/2A inhibition or by PKC activation. PKC inhibition also increased [³H]IP(x) accumulation two- to threefold and changed the Ca²âº response into a peak-plateau response. However, selective inhibition of conventional PKC isoenzymes or preventing changes in [Ca²âº](i) concentration by BAPTA-AM loading was without effect on mGlu5 receptor-stimulated [³H]IP(x) accumulation. Selective knock-down of PKCδ was without effect on glutamate-stimulated Ca²âº responses; however, selective PKCε knock-down in astrocytes changed Ca²âº responses from oscillatory into peak-plateau type. CONCLUSION AND IMPLICATIONS: These data confirm the acute regulation of mGlu5 receptor signalling by protein kinases and protein phosphatases and provide novel data pinpointing the isoenzymic dependence of this regulation in the native mGlu5 receptor-expressing rat cortical astrocyte. These data also highlight a potential alternative mechanism by which mGlu5 receptor signalling might be therapeutically manipulated.

The epidemiology of glycopeptide-resistant enterococci on a haematology unit--analysis by pulsed-field gel electrophoresis.

As part of an interventional study to determine glycopeptide-resistant enterococci (GRE) acquisition on a three-ward haematology unit, rectal swabs were taken weekly from 293 patients recruited to the study between June 1995 and December 1996. The GRE isolates obtained from the first positive rectal swab from 120 colonized patients, the isolates from 7 patients with clinical infection and 43 isolates obtained from the ward environment were compared by pulsed-field gel electrophoresis (PFGE). Sixty-three of 120 patients were colonized by one of strains A-H, while 49 were colonized by unique strains. The first 18 weeks were associated with the highest prevalence of GRE by rectal swab, with a single strain A responsible for 52% of acquisitions on ward 2, 22% on ward 3 and 36% on ward 4. Other smaller ward associated clusters were evident. Environmental sampling of ward 2 during this time showed that all but 2 of 30 isolates were indistinguishable from strain A. As the GRE prevalence fell, rectal swab and environmental isolates became more heterogeneous, and strain A disappeared after week 55. GRE prevalence rose again in the final 15 weeks of the study, and a new predominant strain B emerged on ward 2 responsible for 50% of new acquisitions. In the seven patients with clinical infection with GRE, the clinical isolates were compared with the contemporaneous rectal swab isolate, and were found to be the same in only two cases. An analysis of five long-term carriers colonized for a median of 19 weeks (range 11-34) showed colonization with at least two and in one case six distinct strains, raising the question of how many strains may be colonizing a patient at any one time, and suggesting that multiple colonies should be analysed. These data suggest that cross-infection was an important factor in the spread of GRE when the colonization rate was high.

Handedness in boys with gender identity disorder.

Handedness preference was assessed in 205 boys with gender identity disorder and 205 clinical control boys referred for other reasons. Boys with gender identity disorder were significantly more likely to be left-handed than the clinical control boys (19.5% vs. 8.3%, respectively). The boys with gender identity disorder, but not the clinical control boys, also had a significantly higher rate of left-handedness compared to three independent, general population studies of nonreferred boys (11.8%; N = 14,253) by Hardyck, Goldman, and Petrinovich (1975), Calnan and Richardson (1976), and Eaton, Chipperfield, Ritchot, and Kostiuk (1996). Left-handedness appears to be a behavioral marker of an underlying neurobiological process associated with gender identity disorder in boys.

Synthesis and biological evaluation of sialylmimetics as rotavirus inhibitors.

Rotaviruses cause severe gastroenteritis in infants and are estimated to be responsible for over 600 000 deaths annually, primarily in developing countries. The development of potential inhibitors of this virus is therefore of great interest, particularly since the safety and efficacy of rotaviral vaccines has recently been questioned. This study describes the synthesis of a variety of compounds that can be considered as mimetics of N-acetylneuraminic acid thioglycosides and the subsequent in vitro biological evaluation of these sialylmimetics as inhibitors of rotaviral infection. Our results show that readily accessible carbohydrate-based compounds have the potential to act as inhibitors of rotaviral replication in vitro, presumably through inhibition of the rotaviral adhesion process.

Synthesis of novel sialylmimetics as biological probes.

Glycomimetics are increasingly being recognised as powerful tools in the search for novel compounds that possess useful biological properties. This paper describes our preliminary efforts towards the development of novel mimetics of sialic acid thioglycosides. These sialylmimetics are readily prepared and have been shown, in some instances, to have biological properties similar to sialic acid thioglycosides.

Anxiety and mood disorders in children and adolescents: A practice update.

Anxiety and mood disorders are among the most common disorders in children and adolescents. They presage later emotional difficulties and disabilities. An understanding of the disorders' presentation, common contributing factors and methods of intervention will enable paediatricians and family doctors to provide optimal support to these children and their families. The present paper briefly reviews the epidemiology of anxiety and mood disorders in children and adolescents. Phenomenology is referred to according to the major diagnostic categories for anxiety and depression. Contributing factors, including genetic and environmental components and their possible interaction, are discussed. The management of the disorders, including common strategies for encouraging coping responses, stress reduction and medication, is also described.

Control of glycopeptide-resistant enterococci in an oncology unit.

Enterococci are responsible for 10-12% of nosocomial infections. Since 1987 the incidence of glycopeptide-resistant enterococci (GRE; termed vancomycin-resistant enterococci in the United States) has increased dramatically. The mechanism of GRE is well understood and involves mutation of a single terminal amino acid in a peptidoglycan precursor leading to reduced vancomycin affinity. Studies implicated antibiotic selection pressure as a major risk factor for GRE infection and colonization. In the hematology unit of a London hospital, GRE emerged in December 1993, with 38% of patients positive in a point prevalence study. Between December 1993 and June 1995, 13 patients acquired GRE bacteremia, five of whom died. Between 1995 and 1996 a prospective sequential study was undertaken to determine the effect of changing antibiotic treatment of febrile neutropenia (FN). All patients admitted to the hematology unit were enrolled in the study. In phase 1, treatment continued with ceftazidime monotherapy. In phase 2, piperacillin-tazobactam replaced ceftazidime, and enhanced infection control measures were encouraged. In phase 3, therapy returned to ceftazidime. In phase 1, 57% of patients became colonized or infected with GRE. Phase 2, which was divided into two 4-month cohorts, showed a significant reduction in GRE acquisition, falling to 8% colonization and no clinical infections in the second 4 months. In phase 3, despite heightened infection control procedures, the acquisition rate rose to 36%.

How pure is your thiosialoside? A reinvestigation into the HPLC purification of thioglycosides of N-acetylneuraminic acid.

The synthesis of thiosialosides as potential biological probes for investigations involving the use of sialic acid-recognising proteins has been reinvestigated. It has been found that the most efficient method for the preparation of thiosialosides free from any 2,3-didehydro sialic acid contaminants involves an intermediate HPLC purification of thiosialosides as their methyl esters. Subsequent methyl ester hydrolysis provides thiosialosides (eg. 6 and 14) which are suitable for studies involving the use of sialic acid-recognising proteins.

Nitric oxide synthase inhibition reduces leg glucose uptake but not blood flow during dynamic exercise in humans.

Nitric oxide (NO) appears to play a role in contraction-stimulated glucose uptake in isolated rodent skeletal muscle; however, no studies have examined this question in humans. Seven healthy men completed two 30-min bouts of supine cycling exercise at 60 +/- 2% peak pulmonary oxygen uptake (VO2 peak), separated by 90 min of rest. The NO synthase inhibitor N(G)-monomethyl-L-arginine ([L-NMMA]; total dose 5 mg/kg body weight) or saline (control) were administered via the femoral artery for the final 20 min of exercise in a randomized blinded crossover design. L-Arginine (5 mg/kg body weight) was co-infused during the final 5 min of each exercise bout. Leg blood flow (LBF) was measured by thermodilution in the femoral vein, and leg glucose uptake was calculated as the product of LBF and femoral arteriovenous (AV) glucose difference. L-NMMA infusion significantly (P < 0.05) reduced leg glucose uptake compared with control (48 +/- 12% lower at 15 min, mean +/- SE). The reduction in glucose uptake was due solely to a decrease in AV glucose difference, as there was no effect of L-NMMA infusion on LBF during exercise. Co-infusion of L-arginine restored glucose uptake during L-NMMA infusion to levels similar to control. These results indicate that NO production contributes substantially to exercise-mediated skeletal muscle glucose uptake in humans independent of skeletal muscle blood flow.

Gender constancy judgments in children with gender identity disorder: evidence for a developmental lag.

Gender constancy judgments in children referred for problems in their gender identity development (N = 206) and controls (N = 95) were compared. On Slaby and Frey's (1975) gender constancy interview, the gender-referred children performed more poorly than the controls at three stage levels: gender identity, gender stability, and gender consistency. On the Boy-Girl Identity Task, a second measure of gender constancy (Emmerich et al., 1977), the gender-referred children also performed more poorly. Gender-referred children who had not attained gender consistency engaged in significantly less same-sex-typed play on a free-play task than the gender-referred children who had, but there were no gender consistency effects for the controls. Two other measures of sex-typed behavior were unrelated to gender consistency. In the gender-referred group alone, children who "failed" the gender identity or gender stability stages were more likely to draw an opposite-sex person first on the Draw-a-Person test and to evince more affective gender confusion on the Gender Identity Interview (Zucker et al., 1993) than children who had "passed." It is concluded that children referred for problems in their gender identity development have a developmental lag in gender constancy acquisition. Possible reasons for the lag are discussed.

The control of hyperendemic glycopeptide-resistant Enterococcus spp. on a haematology unit by changing antibiotic usage.

The rectal carriage of glycopeptide-resistant Enterococcus spp. (GRE) had been established at approximately 50% in a series of prevalence studies on a busy haematological malignancy unit. The aim of this study was to reduce the chance of patients acquiring GRE. A prospective three-phase sequential study was performed. In Phase 1, the acquisition rate of GRE detectable by rectal swab was measured without any intervention for a period of 4 months. For the following 8 months (Phase 2), the first-line treatment for febrile neutropenic episodes was changed from monotherapy with ceftazidime to piperacillin/tazobactam. In addition, an intense education programme was introduced to improve hygiene to reduce the risk of case-to-case spread. In the final 4 months (Phase 3), ceftazidime was again used as the first-line antimicrobial, while continuing the same level of training in relation to hygiene. The carriage of GRE was measured from rectal swabs done weekly. During the initial 4 months, at any time, 40-50% of patients in the unit were colonized with GRE, and 43 of 75 (57%) new patients initially negative for GRE acquired it within 6 weeks of their admission. In Phase 2, 25 patients out of 129 (19%) acquired GRE, with the acquisition rate falling progressively so that in the last 3 months, only one new patient acquired GRE (logrank comparison of probabilities for cohort 1 vs cohort 2b: P < 0.0001). A return to ceftazidime in Phase 3 was associated with a return of the risk of acquiring detectable GRE colonization, despite continued hygiene teaching and surveillance, with 21 out of 58 patients (36%) acquiring GRE (cohort 1 vs cohort 3: P = 0.08). Glycopeptide usage was not reduced during the period of the study. Clinical cases were seen only in Phases 1 and 3. Although the reduction in the risk of acquiring GRE may have been due in part to hygiene practices as well as to the change in antimicrobial usage, or may have occurred spontaneously for other reasons, the return of the problem with the reintroduction of ceftazidime strongly suggests that this antibiotic was responsible for encouraging the acquisition of detectable GRE.

Adoptee overrepresentation among clinic-referred boys with gender identity disorder.

OBJECTIVES: To test the hypothesis that adoptees are overrepresented among a sample of clinic-referred boys with gender identity problems (N = 238). To compare the adoptees and nonadoptees on demographic, behaviour problem, and gender-typed measures. METHOD: The percentage of clinic-referred boys with gender identity problems adopted in the first 2 years of life ("early adoptees") was compared to the base rate of boys adopted in Ontario. Parent-report and behavioural measures were used to compare the early adoptees with "late adoptees" (adopted after the second year of life) and nonadoptees. RESULTS: The percentage of boys with gender identity problems who were early adoptees (7.6%) was significantly higher than the base rate of males adopted in Ontario in the first 2 years of life (1.5%). Both the early and late adoptees were significantly less intelligent than the nonadoptees. The early adoptees also had significantly higher externalizing T scores on the Child Behavior Checklist than did the late adoptees and the nonadoptees. The 3 groups did not differ in the percentage who met the complete Diagnostic and Statistical Manual of Mental Disorders (DSM) criteria for gender identity disorder and on 4 other measures of gender-typed behaviour. CONCLUSION: Adoptees are overrepresented among clinic-referred boys with gender identity problems. The reasons for this finding are not clear but may be accounted for by general risk factors that increase the likelihood of clinical referral or by psychosocial and biological factors associated with adoption.

Experiment of nurture: ablatio penis at 2 months, sex reassignment at 7 months, and a psychosexual follow-up in young adulthood.

Guidelines of psychosexual management for infants born with physical intersex conditions are intended to assist physicians and parents in making decisions about sex of assignment and rearing including the following: 1) sex assignment should be to the gender that carries the best prognosis for good reproductive function, good sexual function, normal-looking external genitalia and physical appearance, and a stable gender identity; 2) the decision regarding sex assignment should be made as early as possible, preferably during the newborn period, with an upper age limit for reversal of an initial sex assignment no later than 18 to 24 months; and 3) there should be minimal uncertainty and ambiguity on the part of parents and professionals regarding the final decision about sex assignment and rearing. J. Money used these guidelines in a case of a biologically normal male infant (one of a pair of monozygotic twins) whose penis was accidentally ablated during a circumcision at the age of 7 months. The decision to reassign the infant boy to the female sex and to rear him as a girl was made at 17 months, with surgical castration and initial genital reconstruction occurring at 21 months. Money reported follow-up data on this child through the age of 9 years. Although the girl was described as having many "tomboyish" behavioral traits, a female gender identity had apparently differentiated. Thus, it was concluded that gender identity is sufficiently incompletely differentiated at birth as to permit successful assignment of a genetic male as a girl, in keeping with the experiences of rearing. Subsequent follow-up by other investigators reported that by early adolescence the patient had rejected the female identity and began to live as a male at the age of 14 years. In adulthood, the patient recalled that he had never felt comfortable as a girl, and his mother reported similar recollections. At age 25, the patient married a woman and adopted her children. The patient reported exclusive sexual attraction to females. The present case report is a long-term psychosexual follow-up on a second case of ablatio penis in a 46 XY male. During an electrocautery circumcision at the age of 2 months, the patient sustained a burn of the skin of the entire penile shaft, and the penis eventually sloughed off. At age 7 months, the remainder of the penis and the testes were removed. By age 7 months, if not earlier, the decision was made to reassign the patient as a female and to raise the infant as a girl. The patient was interviewed on two occasions: at 16 years and twice while in the hospital for additional surgery at 26 years of age. At ages 16 and 26, the patient was living socially as a woman and denied any uncertainty about being a female. During childhood, the patient recalled that she self-identified as a "tomboy" and enjoyed stereotypically masculine toys and games; however, the patient also recalled that her favorite playmates were usually girls and that her best friend was always a girl. When seen at age 16, the patient had been admitted to the hospital for vaginoplasty. At that time, she wished to proceed with the further repair of her genitalia to make them suitable for sexual intercourse with males. At age 26, the patient returned to the hospital for further vaginoplasty. Regarding the patient's sexual orientation, she was attracted predominantly to women in fantasy, but had had sexual experiences with both women and men. At the time of the second surgery, she was in a relationship with a man and wished to be able to have intercourse. The patient's self-described sexual identity was "bisexual." After surgery at age 26 years, the patient developed a rectovaginal fistula. Within a few months of the surgery, the patient and her male partner separated for reasons other than the patient's physical problems. The patient subsequently began living with a new partner, a woman, in a lesbian relationship. The psychosexual development of our patient was bot the other patient was married to a woman. Our patient had a "bisexual" sexual identity; the other patient had a "heterosexual" sexual identity. The patients were similar in that they had a childhood history of "tomboyism." Our patient was predominantly sexually attracted to women; the other patient was exclusively sexually attracted to women. Our patient had sexual experiences with both women and men; the other patient had sexual experiences only with women. The most plausible explanation of our patient's differentiation of a female gender identity is that sex of rearing as a female, beginning at around age 7 months, overrode any putative influences of a normal prenatal masculine sexual biology. Because cases of ablatio penis in infancy are so rare and long-term follow-up data are scant, it is obviously impossible to know whether our patient or the previous case would be more typical of the psychosexual outcome in a larger sample of such individuals. However, our case suggests that it is possible for a female gender identity to differentiate in a biologically "normal" genetic male. At present, however, the clinical literature is deeply divided on the best way to manage cases of traumatic loss of the penis during infancy. Further study is clearly required to decide on the optimal model of psychosocial and psychosexual management for affected individuals.

Birth order in girls with gender identity disorder.

This study examined the birth order of girls with gender identity disorder (N = 22). Each proband was matched to 3-7 clinical control girls for age at assessment and number of siblings (the mode number of controls per proband was 7) (total N = 147). The number of older brothers, older sisters, younger brothers, and younger sisters was recorded. Slater's birth order index showed that the probands were significantly more likely to be early born than were the controls. A modified Slater's index also compared the birth order of the probands and the controls only to their brothers (when they had one or more) and only to their sisters (when they had one or more). Compared to the controls, the probands were born early compared to their sisters, but not to their brothers. These findings are the inverse of two previous studies of boys with gender identity disorder, who were later born relative to clinical control boys, an effect that appeared to be accounted for primarily by being born later relative to older brothers, but not to older sisters.

Effects of physical appearance on masculine trait ratings of boys and girls with gender identity disorder.

University students, masked to group status, rated the physical appearance of boys and girls with gender identity disorder (GID) and same-sex controls using traits with stereotypical masculine connotations (for boys: all-boy, handsome, masculine, and rugged; for girls: handsome, masculine, rugged, and tomboyish). Three traits (all-boy, masculine, and rugged) were judged to be significantly less characteristic of the boys with GID as compared to the same-sex controls; for the fourth trait (handsome), the inverse result was obtained. All four traits (handsome, masculine, rugged, and tomboyish) were judged to be significantly more characteristic of the girls with GID as compared to the same-sex normal and/or clinical controls. These results were the inverse of the results obtained in two previous experiments, in which traits with stereotypical feminine connotations were used (Fridell et al., 1996; Zucker et al., 1993). Taken together, the results suggest that boys and girls with GID have a sex-typed physical appearance that distinguishes them from same-sex controls. Possible determinants of these differences are discussed.

Non-transferrin-bound iron induced by myeloablative chemotherapy.

Previous studies have suggested that non-transferrin-bound plasma iron (NTBI) is present in patients undergoing cytotoxic chemotherapy, and that this may exacerbate untoward organ damage and increase the risk of bacterial infections following chemotherapy. However, the source of NTBI during myelosuppressive chemotherapy is controversial. In this study we have examined the kinetics of the appearance and disappearance of NTBI with chemotherapy. NTBI was present in only two out of 24 patients prior to chemotherapy but, following chemotherapy, was present in 19 patients, with peak NTBI levels at 5 d after onset of chemotherapy (mean 3.06 microM). Thereafter levels fell, but were still detectable in seven patients 14 d after the end of chemotherapy. The appearance of NTBI was associated with a sudden rise in transferrin saturation, but NTBI was detected on four occasions in the absence of full transferrin saturation. We have also established that daunorubicin cannot itself liberate NTBI from serum. There was no relationship between induced NTBI levels and serum iron or ferritin levels, previous or current blood transfusions, disease stage, or the type of chemotherapy given. The appearance of NTBI following chemotherapy was inverserely related to the fall in reticulocytes and serum transferrin receptor (TfR) levels, suggesting that NTBI formation is a consequence of suspension of erythropoietic activity.

Gender identity disorder: a review of the past 10 years.

OBJECTIVE: To review the clinically relevant literature on gender identity disorder (GID) in children and adolescents over the past 10 years. METHOD: All literature referring to gender identity and children or adolescents from 1985 on was reviewed. RESULTS: Changes in the DSM-IV nomenclature include (1) adoption of the single diagnosis of GID to apply to children, adolescents, and adults; (2) changes in the format of the criteria; and (3) placement in the section "Sexual and Gender Identity Disorders." Rates of associated psychopathology in children with GID are comparable with those in children with other psychiatric disorders, particularly disorders that are internalizing in form. Biological and psychosocial factors thought to be relevant in the development of GID are reviewed. CONCLUSIONS: Research is required to elucidate the complicated interaction between biological and psychosocial factors in the development of GID and to evaluate treatment efficacy.

Sibling sex ratio of boys with gender identity disorder.

Sibling sex ratio (the ratio of brothers to sisters) was calculated for 444 boys with gender identity disorder (or with behaviors consistent with this diagnosis). The probands were ascertained from several researchers with expertise with this disorder and from the English language case report literature between 1938 and 1995. Among the probands with at least one sibling (N = 333), the results showed that boys with gender identity disorder had a significant excess of brothers to sisters, 131.1:100, when compared with the expected secondary sex ratio of 106:100. The excess of brothers replicated a previous study by Blanchard, Zucker, Bradley, and Hume (1995), in which the sibling sex ratio was 140.6:100. Further analyses showed that the probands were born later relative to their brothers than they were relative to their sisters. These findings are amenable to several psychosocial and biological explanations, which require further investigation.

Sex differences in referral rates of children with gender identity disorder: some hypotheses.

From 1978 through 1995, a sex ratio of 6.6:1 of boys to girls (N = 275) was observed for children referred to a specificity clinic for gender identity disorder. This article attempts to evaluate several hypotheses regarding the marked sex disparity in referral rates. The sexes did not differ on four demographic variables (age at referral, IQ, and parent's social class and marital status) and on five indices of general behavior problems on the Child Behavior Checklist; in addition, there was only equivocal evidence that boys with gender identity disorder had significantly poorer peer relations than girls with gender identity disorder. Although the percentage of boys and girls who met the complete DSM-III-R criteria for gender identity disorder was comparable, other measures of sex-typed behavior showed that the girls had more extreme cross-gender behavior than the boys. Coupled with external evidence that cross-gender behavior is less tolerated in boys than in girls by both peers and adults, it is concluded that social factors partly account for the sex difference in referral rates. Girls appear to require a higher threshold than boys for cross-gender behavior before they are referred for clinical assessment.

Assessment of brain function in adolescent anorexia nervosa before and after weight gain.

This study assessed brain function in 20 adolescent females with anorexia nervosa (AN) and 20 controls using event-related potentials (ERPs) and a battery of neuropsychological tests. In the AN group, N4 latencies for a nonverbal memory task were significantly longer than for a verbal task, and P3 latencies for the verbal task were significantly longer among anorexics as compared to controls. On the nonverbal task, the AN group failed to show a right > left hemispheric asymmetry for P3 amplitudes which was observed for controls. These group differences for P3 latency and amplitude were particularly pronounced in the central-parietal region of the head. Body Mass Index (BMI) in the anorexic group significantly predicted N4 amplitudes for the verbal task in the left hemisphere and P3 amplitudes for the nonverbal task in the right hemisphere. The two groups did not differ on any of the tests used to assess neuropsychological functioning. Eight nutritionally recovered patients and their matched controls were retested using the same procedures. Anorexics showed larger P3 amplitudes for the verbal as compared to the nonverbal task at follow-up. These findings provide evidence for localized brain dysfunction in anorexia nervosa that only partially normalizes with weight gain.