Maturation and fertilization in Lottia gigantea oocytes: intracellular pH, Ca(2+), and electrophysiology.

Intracellular pH and Ca(2+) were measured with BCECF- and Calcium Green-dextran during maturation and fertilization of oocytes of the limpet Lottia gigantea. Maturation of oocytes from prophase to metaphase I of meiosis was induced in seawater adjusted to pH 9 with NH(4)OH. Intracellular pH rose during maturation induction, and maturation was also induced by microinjecting pH 8, but not pH 7, HEPES buffer. Intracellular Ca(2+) rose during NH(4)OH-induced maturation, but maturation was not inhibited when the increase was blocked by microinjection of BAPTA. When the metaphase I oocytes were fertilized(), there was an abrupt increase in intracellular Ca(2+), and activation (polar body formation) failed to occur in BAPTA-injected oocytes. Intracellular pH did not rise during fertilization. These observations show that maturation from prophase to metaphase I of meiosis is pH-dependent and activation of the metaphase I oocytes is Ca(2+)-dependent. A Ca(2+) action potential was present in both immature and mature oocytes but was more prominent in mature oocytes whose input resistance was higher. Fertilization produced a long-lasting (17-20 min) Na(+)-dependent fertilization potential with superimposed oscillations resembling Ca(2+) action potentials.

Psychosocial and risk behavior correlates of youth suicide attempts and suicidal ideation.

OBJECTIVE: To identify the independent psychosocial and risk behavior correlates of suicidal ideation and attempts. METHOD: The relationships between suicidal ideation or attempts and family environment, subject characteristics, and various risk behaviors were examined among 1,285 randomly selected children and adolescents, aged 9 through 17 years, of whom 42 (3.3%) had attempted suicide and 67 (5.2%) had expressed suicidal ideation only. The youths and their parents were enumerated and interviewed between December 1991 and July 1992 as part of the NIMH Methods for the Epidemiology of Child and Adolescent Mental Disorders (MECA) Study. RESULTS: Compared with subjects with suicidal ideation only, attempters were significantly more likely to have experienced stressful life events, to have become sexually active, to have smoked more than one cigarette daily, and to have a history of ever having smoked marijuana. After adjusting for sociodemographic characteristics, a statistically significant association was found between suicidal ideation or attempt and stressful life events, poor family environment, parental psychiatric history, low parental monitoring, low instrumental and social competence, sexual activity, marijuana use, recent drunkenness, current smoking, and physical fighting. Even after further adjusting for the presence of a mood, anxiety, or disruptive disorder, a significant association persisted between suicidal ideation or attempts and poor family environment, low parental monitoring, low youth instrumental competence, sexual activity, recent drunkenness, current smoking, and physical fighting. CONCLUSION: Low parental monitoring and risk behaviors (such as smoking, physical fighting, alcohol intoxication, and sexual activity) are independently associated with increased risk of suicidal ideation and attempts, even after adjusting for the presence of psychiatric disorder and sociodemographic variables.

Risk behavior in a community sample of children and adolescents.

OBJECTIVES: First, to investigate whether there is covariation between risk behaviors, including suicidality, in a community probability sample of children and adolescents; and second, to investigate whether risk behavior is associated with selected potential correlates. METHOD: A sample of 9- to 17-year-old youths (N = 1,285) and their caretakers were interviewed in the Methods for the Epidemiology of Child and Adolescent Mental Disorders (MECA) Study. The risk behaviors were marijuana smoking, alcohol use, intercourse, fighting, cigarette smoking, and suicidal ideation/attempts. Relationships between the risk behaviors were described using odds ratios. Linear regression analyses of an index of risk behavior on the selected potential correlates of risk behavior were conducted. RESULTS: There were significant relationships between all pairs of risk behaviors. The score on the index of risk behavior was associated with stressors, lack of resources, family psychiatric disorder, psychopathology, and functional impairment. CONCLUSIONS: Clinicians should be alerted to the possibility of risk behaviors, especially in children and adolescents engaging in other risk behaviors and those with inadequate resources, stressors, functional impairment, or psychopathology.

MAP kinase, a universal suppressor of sperm centrosomes during meiosis?

We reported previously that inhibition of MAP kinase during meiosis in Urechis caupo eggs caused premature sperm aster formation and we reviewed indirect evidence that the suppression of sperm asters by MAPK during meiosis might be a universal mechanism (M. C. Gould and J. L. Stephano, 1999, Dev. Biol. 216, 348-358). We tested this proposition with oyster (Crassostrea gigas) and starfish (Asterina miniata) eggs, utilizing the MEK inhibitors U0126 and PD98059. Centrosomes, asters, and meiotic spindles were visualized by normal epifluorescence and confocal microscopy following indirect immunocytochemical staining for anti-beta-tubulin. When MAPK activation was inhibited, sperm asters in both species developed prematurely and tended to move toward the egg centrosomes, sometimes even fusing with the egg spindle or centrosomes. Meiotic spindles and polar body formation were also abnormal when MAPK was inhibited.

MAP kinase, meiosis, and sperm centrosome suppression in Urechis caupo.

Although MAP kinase is an important regulatory enzyme in many somatic cells, almost nothing is known about its functions during meiosis, except in frog and mouse oocytes. We investigated MAPK activation and function in oocytes of the marine worm Urechis caupo that are fertilized at meiotic prophase. Activity was first detected at 4-6 min after fertilization in immunoblots with anti-active MAPK, prior to germinal vesicle breakdown (GVBD). MAPK activation did not require new protein synthesis and was dependent on the increases in both intracellular pH and intracellular Ca(2+) that normally occur during activation. When MAPK activation was inhibited with PD98059 or U0126, GVBD still occurred, but meiosis was abnormal and there was a dramatic premature enlargement of sperm asters, which normally do not appear until second polar body formation. Failure of polar body formation and premature sperm aster enlargement also occurred when MAPK activation was inhibited by an entirely different treatment which involved lowering the pH of external seawater to interrupt the normal cytoplasmic pH increase. Thus, in Urechis, active MAPK appears to be required for (1) normal meiotic divisions and (2) suppressing the paternal centrosome until after the egg completes meiosis, a general phenomenon whose mechanism has been unknown.

Parthenogenesis in Urechis caupo (Echiura). I. Persistance of functional maternal asters following activation without meiosis.

A peptide (P23) isolated from sperm acrosomal protein initiates development in eggs of the marine worm Urechis caupo (Echiura). However, in these earlier experiments the eggs were exposed continuously to P23 and did not cleave unless they were provided with a sperm centrosome. The present study reports that parthenogenetic cleavage can be induced when the eggs are exposed to P23 for 1.5 to 2 min in seawater at pH 8 followed by acidifying the seawater to pH 7 or washing off the peptide at pH 7 or 8. The mechanism of parthenogenesis induction was studied by comparing the behavior of maternal and paternal centrosomes (microtubule organizing centers), chromosomes, nuclei and nucleoli in fertilized eggs, P23-activated eggs that failed to cleave, and P23-activated eggs that cleaved, using bright-field, phase-contrast, and fluorescence microscopy following anti-tubulin and bis-benzimide staining. Parthenogenetic cleavage to mostly diploid embryos occurred in eggs that underwent germinal vesicle breakdown without polar body formation. The centrosomes used for cleavage were the persistent maternal centrosomes that did not cycle through meiosis. When cytochalasin B was used to suppress polar body formation without inhibiting the meiotic cycles, eggs did not subsequently cleave even though all four maternal centrosomes were retained. When fertilized eggs were transferred to pH 7 seawater at 1.5 to 2 min after insemination, they underwent a partial activation like the P23-activated eggs and did not form polar bodies. Subsequent cleavage was very abnormal due to the presence of multiple asters formed by the persistant active maternal centrosomes. These results show that maternal centrosomes are normally inactivated by a maternal mechanism associated with meiotic cycling.

Nuclear and cytoplasmic pH increase at fertilization in Urechis caupo.

Intracellular pH (pHi) was measured in Urechis caupo (Echiura) eggs during fertilization using the pH-sensitive dye BCECF [bis(carboxyethyl)carboxyfluorescein] and fluorescence microscopy. When eggs were inseminated at pH 8, pHi began to rise 22-36 sec (n = 7) after sperm contact and reached a plateau by 3 min (2.8 +/- 1 SD; n = 14). The net increased was 0.25-0.3 pH units and the alkalinization persisted through 1 hr after insemination (after second polar body formation). Separate measurements of germinal vesicles and cytoplasm revealed that pH rose dramatically within the nuclei well before germinal vesicle breakdown (GVBD), as well as in cytoplasm. When eggs were fertilized at pH 6.9 (previously shown to inhibit proton release and egg activation despite sperm entry; see Paul, 1970, 1975; Holland et al., 1984), there was no net pH increase in either cytoplasm or germinal vesicles. When fertilization was at pH 7.45, the average pHi increase in whole eggs was 58% of that at pH 8, and 61% of the eggs activated partially or completely. These results show that the pHi rise is correlated with GVBD and egg activation.

Correspondence between statistically derived behavior problem syndromes and child psychiatric diagnoses in a community sample.

The correspondence between Diagnostic and Statistical Manual (3rd ed.) (DSM-III) diagnoses and statistically derived syndromes was examined within a community sample of children and adolescents in Puerto Rico. Specifically, the extent to which behavior dimensions, derived from the Child Behavior Checklist and the Youth Self-Report, corresponded to psychiatric diagnoses, derived from parent and child versions of the Diagnostic Interview Schedule for Children, was examined. The alternative approaches for assessing psychopathology in children and adolescents were compared against external validators. The results indicated a meaningful convergence between DSM-III diagnoses and statistical syndromes; however, a one-to-one correspondence did not emerge. Little evidence was found for "diagnostic thresholds." There was no evidence of the superiority of either the statistically derived syndromes or the DSM-III diagnoses. The incorporation of a measure of impairment improved the validity of both approaches. Adding parental reports to the self-reports of adolescents yielded little gain in the validity of either the statistical or diagnostic approach. The implications for the definition and assessment of child and adolescent disorders are discussed.

Patterns of diagnostic comorbidity in a community sample of children aged 9 through 16 years.

Secondary analyses of the data from the Puerto Rico Child Psychiatry Epidemiologic Study were done to provide information on the comorbidity of four major diagnostic domains (attention deficit disorders, conduct/oppositional disorders, depression and anxiety disorders). A high level of comorbidity was found among these four domains of child and adolescent psychopathology. In general the patterns of comorbidity were not affected by whether the data was put together by a clinician or by means of a computer algorithm scoring a structured interview. The patterns were not affected in any major way by who the informants were in the diagnostic process. Minor differences were found in certain comorbidity patterns depending on the sex and age of the subjects. Comorbidity was associated with level of impairment and to service utilization.

Aggregating data from multiple informants in child psychiatry epidemiological research.

One of the issues confronting the field of child psychiatric epidemiology is devising ways to aggregate discrepant reports from multiple informants obtained in structured diagnostic assessments of children and adolescents. The present report uses data from an epidemiological community survey to attempt to address this issue. The results obtained through statistical procedures designed to identify the "optimal" informant for specific symptomatic criteria and diagnoses do not seem to offer a clear-cut advantage over a simple combinatorial rule that identifies symptomatic criteria as positive when they are acknowledged as positive by either parent or child informants.

Screening for childhood psychopathology in the community using the Child Behavior Checklist.

One of the uses of the Child Behavior Checklist is as a screening instrument for childhood psychopathology in two-phase designs. The present report involves a two-phase epidemiological survey conducted in Puerto Rico in which the CBCL was used as a screening instrument during the first stage, and children were evaluated clinically during the second stage. The data indicate that in using the CBCL for screening for psychopathology in children, parent information is most informative, particularly for children in the adolescent age group. Nevertheless, the data also reinforce the need to obtain teacher information with the Teacher Report Form to enhance screening sensitivity. In this population, the data obtained with the Youth Self-Report were found to be of limited usefulness for screening purposes.

Impairment in the epidemiological measurement of childhood psychopathology in the community.

The desirability of incorporating a measure of impairment to the categorization of childhood psychopathology in the community is examined. The use of the Children's Global Assessment Scale (CGAS) for this purpose is recommended. The choice of 61 (definite case) and 71 (probable case) as cutpoints on the Children's Global Assessment Scale is supported empirically by the data on service utilization, parental perceived need, and behavior problem scores obtained in the Puerto Rico Child Psychiatry Epidemiological Study.

The internal consistency and concurrent validity of a Spanish translation of the Child Behavior Checklist.

The Child Behavior Checklist's applicability to a Hispanic sample was assessed by an examination of the instrument's internal consistency and concurrent validity. The CBCL and TRF were administered to a community sample representative of children of Puerto Rico aged 4 to 16. Cronbach's alpha was used to assess the internal consistency of empirically derived scales. The relation of CBCL and TRF scores to clinical diagnosis, adaptive functioning, and need for services served as indicators of the concurrent validity of the instrument's Spanish version. The results indicate that the total behavior problem scores on the instruments are good continuous measures of maladjustment for children in Puerto Rico. A child with high values on the scales has a high probability of being classified as a case by a psychiatrist. High levels of internal consistency were found in most subscales. Only scales comprising low prevalence problems showed poor internal consistency.

Epidemiological comparisons of Puerto Rican and U.S. mainland children: parent, teacher, and self-reports.

U.S. mainland and Puerto Rican nonreferred samples were compared via the Child Behavior Checklist (ages 4 to 16), Teacher's Report Form (ages 6 to 16), and Youth Self-Report (ages 12 to 16). Problem scores were significantly higher in parent and teacher ratings of Puerto Rican than mainland subjects, but were significantly lower in self-ratings by Puerto Rican adolescents. Adolescents in both cultures reported significantly more problems than their parents or teachers did. Most of the significant cross-cultural differences in parent, teacher, and self-ratings of competencies showed more favorable scores for the mainland subjects. High referral rates, a high prevalence of DSM diagnoses, and low scores on the Children's Global Assessment Scale are consistent with the high problem rates reported by Puerto Rican parents and teachers but not with the lower rates reported by adolescents. Different clinical cutoffs may be needed for all assessments in the mainland versus Puerto Rico.

Risk factors for maladjustment in Puerto Rican children.

Data are presented on risk factors for childhood psychopathology derived from a study of an island-wide probability sample of children in Puerto Rico aged 4 through 16 years. Analyses estimated the effects of 12 demographic, health, and family variables on the probability of being a "case," using two different operational definitions of caseness, as well as on the probability of receiving the diagnoses of oppositional disorder, attention deficit disorder, separation anxiety, depression, functional enuresis, and adjustment disorder. When compared to other findings, the results from these analyses indicate that the relationship between maladjustment and the risk factors evaluated does not appear to be culturally specific.

Estimates of the prevalence of childhood maladjustment in a community survey in Puerto Rico. The use of combined measures.

A two-stage epidemiologic survey was carried out on a probability sample of the population aged 4 through 16 years in Puerto Rico. The survey used the Child Behavior Checklist as a screening instrument, and prevalence rates were estimated on the basis of clinical diagnoses and other measures provided by child psychiatrists during the second stage. Maladjustment was operationally defined through the use of combined measures, including DSM-III diagnosis and a scale of functional impairment. Data were provided on the demographic correlates of maladjustment and on the comorbidity of DSM-III diagnostic domains. The prevalence rates obtained vis-à-vis the availability of mental health services on the island reflected a major public health problem.