Using pictures to evoke spiritual feelings in breast cancer patients: development of a new paradigm for neuroimaging studies.

This study was designed to develop and validate a method for enhancing spiritual feelings, particularly in women who have received a diagnosis of breast cancer. The protocol specifically was developed to be used in functional magnetic resonance imaging (fMRI) studies. Eighteen breast cancer survivors rated pictures for their ability to enhance feelings of spirituality, happiness, and sadness. Results indicate that presenting carefully selected pictures with spiritual content (e.g., nature scenes, people engaged in contemplative behaviors) can effectively enhance spiritual feelings among breast cancer survivors. Future fMRI studies will explore the use of the protocol developed in this study for investigating neural activity during spiritual feelings and states.

Cosmogenic chlorine-36 chronology for glacial deposits at bloody canyon, eastern sierra nevada.

Deposits from mountain glaciers provide an important record of Quaternary climatic fluctuations but have proved difficult to date directly. A chronology has been obtained for glacial deposits at Bloody Canyon, California, by measurement ofthe accumulation of chlorine-36 produced by cosmic rays in boulders exposed on moraine crests. The accumulation ofchlorine-36 indicates that episodes of glaciation occurred at about 21, 24, 65, 115, 145, and 200 ka (thousand years ago). Although the timing of the glaciations correlates well with peaks of global ice volume inferred from the marine oxygen isotope record, the relative magnitudes differ markedly. The lengths of the moraines dating from 115 ka and 65 ka show that the early glacial episodes were more extensive than those during the later Wisconsin and indicate that the transition from interglacial to full glacial conditions was rapid.

Protocol: Using N-of-1 tests to identify responders to melatonin for sleep disturbance in Parkinson's disease.

BACKGROUND: 40% of Parkinson's Disease (PD) sufferers experience insomnia, impacting health and quality of life for patients and family members, especially carers. There is little evidence that current treatments are effective. OBJECTIVES: To determine the effectiveness of melatonin in reducing insomnia in 44 individuals with PD using N-of-1 trials. To aggregate group data to arrive at population estimates of effectiveness (measured by improvements in PDSS-2) and safety (measured by adverse events) of melatonin in improving insomnia in PD. To assess the feasibility of offering N-of-1 trials for insomnia in PD. METHODOLOGY: Participants will receive either immediate-release melatonin or placebo in random order in 3 paired two-week treatment periods (12 weeks total). Based on their response in a two-week run-in period on 3 mg daily, they will trial either 3 mg or 6 mg. Patients will keep daily sleep diaries and wear a MotionWatch throughout. After the trial patients will discuss their individual report with their doctor, which provides direct feedback about effectiveness and safety of melatonin for them. STATISTICAL METHODS: We will analyse N-of-1 tests 1) individually: effects of melatonin on PDSS-2 and safety will be reported; and 2) aggregated across individual N-of-1 studies, combined using a Bayesian multilevel random effects model, which will account for repeated measures on individuals over time, and will return posterior estimates of overall treatment effect, and effect in each individual. CLINICAL TRIAL REGISTRATION NUMBER: ACTRN12617001103358.

Nanotechnology of emerging targeting systems.

Recent developments in the design and testing of complex nanoscale payload-carrying systems (i.e. systems with payloads that do not exceed 100 nm in size) are the focus of this brief review. Emerging systems include targeted single-walled nanotubes, viral capsids, dendrimers, gold nanoparticles, milled boron carbide nanoparticles, and protein nucleic acid assemblies. Significant advances are emerging with each of these bionanotechnological approaches to cellular targeting.

Sleep-related vehicle crashes on low speed roads.

BACKGROUND: Very little is known about the characteristics of sleep related (SR) crashes occurring on low speed roads compared with current understanding of the role of sleep in crashes occurring on high speed roads e.g. motorways. To address this gap, analyses were undertaken to identify the differences and similarities between (1) SR crashes occurring on roads with low (≤60km/h) and high (≥100km/h) speed limits, and (2) SR crashes and not-SR crashes occurring on roads with low speed limits. METHOD: Police reports of all crashes occurring on low and high speed roads over a ten year period between 2000 and 2009 were examined for Queensland, Australia. Attending police officers identified all crash attributes, including 'fatigue/fell asleep', which indicates that the police believe the crash to have a causal factor relating to falling asleep, sleepiness due to sleep loss, time of day, or fatigue. Driver or rider involvement in crashes was classified as SR or not-SR. All crash-associated variables were compared using Chi-square tests (Cramer's V=effect size). A series of logistic regression was performed, with driver and crash characteristics as predictors of crash category. A conservative alpha level of 0.001 determined statistical significance. RESULTS: There were 440,855 drivers or riders involved in a crash during this time; 6923 (1.6%) were attributed as SR. SR crashes on low speed roads have similar characteristics to those on high speed roads with young (16-24y) males consistently over represented. SR crashes on low speed roads are noticeably different to not-SR crashes in the same speed zone in that male and young novice drivers are over represented and outcomes are more severe. Of all the SR crashes identified, 41% occurred on low speed roads. CONCLUSION: SR crashes are not confined to high speed roads. Low speed SR crashes warrant specific investigation because they occur in densely populated areas, exposing a greater number of people to risk and have more severe outcomes than not-SR crashes on the same low speed roads.

Sleep-related crash characteristics: Implications for applying a fatigue definition to crash reports.

Sleep-related (SR) crashes are an endemic problem the world over. However, police officers report difficulties in identifying sleepiness as a crash contributing factor. One approach to improving the sensitivity of SR crash identification is by applying a proxy definition post hoc to crash reports. To identify the prominent characteristics of SR crashes and highlight the influence of proxy definitions, ten years of Queensland (Australia) police reports of crashes occurring in ≥100km/h speed zones were analysed. In Queensland, two approaches are routinely taken to identifying SR crashes. First, attending police officers identify crash causal factors; one possible option is 'fatigue/fell asleep'. Second, a proxy definition is applied to all crash reports. Those meeting the definition are considered SR and added to the police-reported SR crashes. Of the 65,204 vehicle operators involved in crashes 3449 were police-reported as SR. Analyses of these data found that male drivers aged 16-24 years within the first two years of unsupervised driving were most likely to have a SR crash. Collision with a stationary object was more likely in SR than in not-SR crashes. Using the proxy definition 9739 (14.9%) crashes were classified as SR. Using the proxy definition removes the findings that SR crashes are more likely to involve males and be of high severity. Additionally, proxy defined SR crashes are no less likely at intersections than not-SR crashes. When interpreting crash data it is important to understand the implications of SR identification because strategies aimed at reducing the road toll are informed by such data. Without the correct interpretation, funding could be misdirected. Improving sleepiness identification should be a priority in terms of both improvement to police and proxy reporting.

Weighing in on international growth standards: testing the case in Australian preschool children.

Overweight and obesity in preschool-aged children are major health concerns. Accurate and reliable estimates of prevalence are necessary to direct public health and clinical interventions. There are currently three international growth standards used to determine prevalence of overweight and obesity, each using different methodologies: Center for Disease Control (CDC), World Health Organization (WHO) and International Obesity Task Force (IOTF). Adoption and use of each method were examined through a systematic review of Australian population studies (2006-2017). For this period, systematically identified population studies (N = 20) reported prevalence of overweight and obesity ranging between 15 and 38% with most (n = 16) applying the IOTF standards. To demonstrate the differences in prevalence estimates yielded by the IOTF in comparison to the WHO and CDC standards, methods were applied to a sample of N = 1,926 Australian children, aged 3-5 years. As expected, the three standards yielded significantly different estimates when applied to this single population. Prevalence of overweight/obesity was WHO - 9.3%, IOTF - 21.7% and CDC - 33.1%. Judicious selection of growth standards, taking account of their underpinning methodologies and provisions of access to study data sets to allow prevalence comparisons, is recommended.

Structural organization of interspersed repetitive elements present in the DNA of Mus musculus.

Two-dimensional displays of the restriction fragments from the DNA of Mus musculus revealed a complex species-specific pattern produced from nonsatellite repetitive sequences. The patterns have been used as a guide in the direct purification of a group of broadly interspersed repeated DNA sequences (characterized by a 1350-bp Eco-Bam fragment) that have been studied by molecular cloning, restriction mapping and genomic Southern blotting. These studies show that the cloned representatives originate from an abundant group of sequences that share homology with about 2% of the mouse genome. The sequences do not appear to share homology with mouse-interspersed-family-1 (MIF-1) nor with the major AT-rich satellite sequences of mouse. They appear to be part of a group of larger repetitive elements that is both broadly interspersed and heavily methylated in normal mouse tissue.

Reversal of somatostatin inhibition of insulin and glucagon secretion.

These studies were designed to elucidate the mechanism of inhibitory action of somatostatin (SRIF) on glucagon (IRG) and insulin (IRI) secretion. Studies were carried out in the unrecirculated isolated rat pancreas perfusion with arginine 19.2 mM and glucose 5.5 mM as stimulus primarily for IRG but also IRI secretion. The effects of excess Ca++ (15.2 mEq./L.) and excess K+ (12.8 mEq./L.) on IRG, IRI, and the SRIF-inhibited pancreas were studied. Ca++ excess in five perfusions strikingly stimulated IRG secretion (+92 per cent) but only stabilized IRI secretion compared with control perfusions. K+ excess (in seven perfusions) markedly inhibited IRG secretion (-39 per cent) while stimulating IRI secretion (+16 per cent). Restoration of normal concentrations of K+ resulted in a rebound of IRG to levels 120 per cent that of controls. SRIF, at concentrations from 0.1-20 ng./ml., produced inhibition of both IRG and IRI. In 11 perfusions, with SRIF at 10 ng./ml., IRG decreased more than IRI (-75.2 per cent IRG and -46.9 per cent IRI). In five perfusions, addition of Ca++ (15.2 mEq./L.) 10 minutes after SRIF was started resulted in a reversal of IRG inhibition to 69.4 per cent and IRI to 73.2 per cent of the arginine controls. The reversal by Ca++ of SRIF effect on IRG was greater at higher concentrations of Ca++, suggesting some form of competition. In four perfusions, excess K+ reversed SRIF-induced IRI inhibition to 79.6 per cent that of controls but had no effect on IRG inhibition. Studies in vitro with isolated islets revealed that SRIF (2 mug./ml.) inhibited 45Ca uptake of islets as did epinephrine (10(-5) M). It was concluded that SRIF-induced inhibition of hormone release appears related to an action on Ca++ uptake.

Cystic fibrosis transmembrane conductance regulator. Physical basis for lyotropic anion selectivity patterns.

The cystic fibrosis transmembrane conductance regulator (CFTR) Cl channel exhibits lyotropic anion selectivity. Anions that are more readily dehydrated than Cl exhibit permeability ratios (P(S)/P(Cl)) greater than unity and also bind more tightly in the channel. We compared the selectivity of CFTR to that of a synthetic anion-selective membrane [poly(vinyl chloride)-tridodecylmethylammonium chloride; PVC-TDMAC] for which the nature of the physical process that governs the anion-selective response is more readily apparent. The permeability and binding selectivity patterns of CFTR differed only by a multiplicative constant from that of the PVC-TDMAC membrane; and a continuum electrostatic model suggested that both patterns could be understood in terms of the differences in the relative stabilization of anions by water and the polarizable interior of the channel or synthetic membrane. The calculated energies of anion-channel interaction, derived from measurements of either permeability or binding, varied as a linear function of inverse ionic radius (1/r), as expected from a Born-type model of ion charging in a medium characterized by an effective dielectric constant of 19. The model predicts that large anions, like SCN, although they experience weaker interactions (relative to Cl) with water and also with the channel, are more permeant than Cl because anion-water energy is a steeper function of 1/r than is the anion-channel energy. These large anions also bind more tightly for the same reason: the reduced energy of hydration allows the net transfer energy (the well depth) to be more negative. This simple selectivity mechanism that governs permeability and binding acts to optimize the function of CFTR as a Cl filter. Anions that are smaller (more difficult to dehydrate) than Cl are energetically retarded from entering the channel, while the larger (more readily dehydrated) anions are retarded in their passage by "sticking" within the channel.

CFTR: covalent modification of cysteine-substituted channels expressed in Xenopus oocytes shows that activation is due to the opening of channels resident in the plasma membrane.

Some studies of CFTR imply that channel activation can be explained by an increase in open probability (P(o)), whereas others suggest that activation involves an increase in the number of CFTR channels (N) in the plasma membrane. Using two-electrode voltage clamp, we tested for changes in N associated with activation of CFTR in Xenopus oocytes using a cysteine-substituted construct (R334C CFTR) that can be modified by externally applied, impermeant thiol reagents like [2-(trimethylammonium)ethyl] methanethiosulfonate bromide (MTSET+). Covalent modification of R334C CFTR with MTSET+ doubled the conductance and changed the I-V relation from inward rectifying to linear and was completely reversed by 2-mercaptoethanol (2-ME). Thus, labeled and unlabeled channels could be differentiated by noting the percent decrease in conductance brought about by exposure to 2-ME. When oocytes were briefly (20 s) exposed to MTSET+ before CFTR activation, the subsequently activated conductance was characteristic of labeled R334C CFTR, indicating that the entire pool of CFTR channels activated by cAMP was accessible to MTSET+. The addition of unlabeled, newly synthesized channels to the plasma membrane could be monitored on-line during the time when the rate of addition was most rapid after cRNA injection. The addition of new channels could be detected as early as 5 h after cRNA injection, occurred with a half time of approximately 24-48 h, and was disrupted by exposing oocytes to Brefeldin A, whereas activation of R334C CFTR by cAMP occurred with a half time of tens of minutes, and did not appear to involve the addition of new channels to the plasma membrane. These findings demonstrate that in Xenopus oocytes, the major mechanism of CFTR activation by cAMP is by means of an increase in the open probability of CFTR channels.

CFTR: covalent and noncovalent modification suggests a role for fixed charges in anion conduction.

The goal of the experiments described here was to explore the possible role of fixed charges in determining the conduction properties of CFTR. We focused on transmembrane segment 6 (TM6) which contains four basic residues (R334, K335, R347, and R352) that would be predicted, on the basis of their positions in the primary structure, to span TM6 from near the extracellular (R334, K335) to near the intracellular (R347, R352) end. Cysteines substituted at positions 334 and 335 were readily accessible to thiol reagents, whereas those at positions 347 and 352 were either not accessible or lacked significant functional consequences when modified. The charge at positions 334 and 335 was an important determinant of CFTR channel function. Charge changes at position 334--brought about by covalent modification of engineered cysteine residues, pH titration of cysteine and histidine residues, and amino acid substitution--produced similar effects on macroscopic conductance and the shape of the I-V plot. The effect of charge changes at position 334 on conduction properties could be described by electrodiffusion or rate-theory models in which the charge on this residue lies in an external vestibule of the pore where it functions to increase the concentration of Cl adjacent to the rate-limiting portion of the conduction path. Covalent modification of R334C CFTR increased single-channel conductance determined in detached patches, but did not alter open probability. The results are consistent with the hypothesis that in wild-type CFTR, R334 occupies a position where its charge can influence the distribution of anions near the mouth of the pore.

Gilbert's conjecture: the search for DNA (cytosine-5) demethylases and the emergence of new functions for eukaryotic DNA (cytosine-5) methyltransferases.

In 1985 Walter Gilbert challenged members of the DNA methylation community assembled at a National Institutes of Health meeting organized by Giulio Cantoni and Ahron Razin with the following words: "The most exciting aspect about the methyl groups on DNA is the thought that they might provide a locally inherited change in a DNA structure. However, for that to be interesting, those changes have to be different in different cells. Furthermore, the alterations in methylation have to be freely imposable and have to be maintained. It is not yet clear that all these properties are true. So I don't think one will find that methylation ever is one of the primary, top-level controls on gene expression."In essence, Gilbert's conjecture, that DNA methylation is not one of the top-level controls on gene expression, assumes that evidence in favor of both of its testable propositions will not be obtained. Evidence for the first proposition, that alterations in methylation status associated with gene-expression states have to be maintained, was already available in 1985 and has been strengthened by a number of very recent experiments. However, the extensive effort to obtain evidence for the second proposition, that alterations in methylation status be freely imposable, has not been successful in its original intent. The effort has, on the other hand, resulted in the emergence of new functions for 5-methylcytosine and the cytosine methyltransferases in eukaryotic DNA repair, recombination and chromosome stability.

Stalling of human DNA (cytosine-5) methyltransferase at single-strand conformers from a site of dynamic mutation.

Single-strand conformers (SSCs) from the C-rich strand of the triplet repeat at the FMR-1 locus are rapidly and selectively methylated by the human DNA (cytosine-5) methyltransferase. The apparent affinity of the enzyme for the FMR-1 SSC is about tenfold higher than it is for a control Watson-Crick paired duplex. The de novo methylation rate for the SSC is over 150-fold higher than the de novo rate for the control duplex. Methylation of what is generally called a hemi-methylated duplex occurs with a rate enhancement of over 100-fold, while methylation of what can be viewed as a hemi-methylated FMR-1 SSC is actually slower than the de novo rate. The pronounced inhibition of the methyltransferase by the methylated SSC suggests that the enzyme has a higher affinity for the methylated product of its reaction with the SSC than it has for the unmethylated SSC substrate. Gel retardation studies show that the methyltransferase binds selectively to SSCs from the C-rich strand of the FMR-1 triplet repeat. This suggests a two-step stalling process in which the human methyltransferase first selectively methlyates and subsequently stalls at the C-rich strand SSC. Stalling may reflect the inability of the enzyme to release a DNA product that is fixed in a conformation resembling its transition state by the unusual structure of the substrate. In particular, the data suggest that DNA methyltransferase may physically participate in biological processes that lead to dynamic mutation at FMR-1. In general, the data raise the possibility that a two-step stalling process occurs at secondary structures associated with chromosome instability, chromosome remodelling, viral replication or viral integration and may account for the local hypermethylation and global hypomethylation associated with viral and non-viral tumorigenesis.

Recognition of unusual DNA structures by human DNA (cytosine-5)methyltransferase.

The symmetry of the responses of the human DNA (cytosine-5)methyltransferase to alternative placements of 5-methylcytosine in model oligodeoxynucleotide duplexes containing unusual structures has been examined. The results of these experiments more clearly define the DNA recognition specificity of the enzyme. A simple three-nucleotide recognition motif within the CG dinucleotide pair can be identified in each enzymatically methylated duplex. The data can be summarized by numbering the four nucleotides in the dinucleotide pair thus: 1 4/2 3. With reference to this numbering scheme, position 1 can be occupied by cytosine or 5-methylcytosine; position 2 can be occupied by guanosine or inosine; position 3, the site of enzymatic methylation, can be occupied only by cytosine; and position 4 can be occupied by guanosine, inosine, O6-methylguanosine, cytosine, adenosine, an abasic site, or the 3' hydroxyl group at the end of a gapped molecule. Replacing the guanosine normally found at position 4 with any of the moieties introduces unusual (non-Watson-Crick) pairing at position 3 and generally enhances methylation of the cytosine at that site. The exceptional facility of the enzyme in actively methylating unusual DNA structures suggests that the evolution of the DNA methyltransferase, and perhaps DNA methylation itself, may be linked to the biological occurrence of unusual DNA structures.

Hypermethylation of telomere-like foldbacks at codon 12 of the human c-Ha-ras gene and the trinucleotide repeat of the FMR-1 gene of fragile X.

Runs of G residues on the G-rich strands of 30mers from the region spanning codon 12 of c-Ha-ras appear to be protected against chemical modification by dimethylsulfate. This suggests that the G-rich strand might spontaneously form a Hoogsteen-paired quadruplex, which is characteristic of telomere-like DNA sequences. In this report we show that the predominant species in 1:1 mixtures of complementary 30mers from this region are duplex DNA and a smaller amount of unimolecular foldback formed by the C-rich strand. Foldbacks of this type resemble structures first observed in the C-rich strand of telomeric DNA and also occur at the CCG triplet repeat present in the FMR-1 gene of human fragile X syndrome. Foldbacks from the C-rich strand of c-Ha-ras and the FMR-1 triplet repeat are exceptional substrates for the human methyltransferase in isolation. Substituting inosine for guanosine alters the secondary structure of the folded oligomers and dramatically reduces their ability to serve as substrates for the human methyltransferase, suggesting that secondary structure is required for recognition by the enzyme. These findings suggest that one mechanism by which methyl groups accumulate in the c-Ha-ras region of chromosome 11 during carcinogenesis and at the FMR-1 locus during repeat expansion at fragile X may be structurally induced de novo methylation at sites undergoing local conformational change. Such methylation might serve to mark unusual structures for repair. In the absence of repair, asymmetrically methylated duplexes produced by resolution of the unusual structures would be rapidly converted to symmetrically methylated duplexes through the methyl-directed activity also carried by the human methyltransferase.

Genetic vulnerability to drug abuse. The D2 dopamine receptor Taq I B1 restriction fragment length polymorphism appears more frequently in polysubstance abusers.

Alcoholics are more likely than nonalcoholics to display the Taq I A1 restriction fragment length polymorphism of the D2 dopamine receptor gene, according to four of six studies that examined alcoholics and controls. The current study examines whether the association observed in alcoholism might extend to other addictive substances by examining D2 dopamine receptor Taq I A and B restriction fragment length polymorphisms in polysubstance users and controls free of significant substance use. We hypothesized a stronger association for the B1 restriction fragment length polymorphism since it lies closer to dopamine receptor protein coding and 5' regulatory regions. Heavy polysubstance users and subjects with DSM-III-R psychoactive substance use diagnoses displayed significantly higher Taq I B1 frequencies than control subjects; Taq I A1 results for these comparisons were less robust. These results are consistent with a role for a D2 dopamine receptor gene variant marked by these restriction fragment length polymorphisms in enhanced substance abuse vulnerability.

Maternal modulation of infantile ovarian development and available ovarian luteinizing hormone-releasing hormone (LHRH) receptors via milk LHRH.

In the rat, ovarian LHRH receptor content, measured by the binding of [125I]D-Ala6-Pro9-LHRH ([125I]A-LHRH) to ovarian membranes, declines during the days preceding the first preovulatory LH surge. The present results show that LHRH receptor content is low during neonatal-infantile days (days 5-15) and increases markedly thereafter to a maximum by day 25. When ovarian membranes from 10-day-old rats were pretreated with MgCl2 to dissociate endogenously bound hormone, LHRH-binding capacity increased to levels similar to those in untreated ovaries from 25-day-old rats. No significant increase was observed in ovaries from the latter group after MgCl2 exposure. Equilibrium association constants were similar (6-8 X 10(9) M-1) in MgCl2-treated and control ovaries, indicating that the increase in binding was not due to a change in receptor affinity. Pups prevented from suckling for various time intervals (1-24 h) exhibited an increase in available ovarian LHRH-binding sites which was maximal by 4 h. When pups were allowed to suckle subsequent to a 6-h fast, available ovarian LHRH receptors decreased rapidly (1 h). This decline was not due to a local increase in LHRH triggered by a neural reflex in response to suckling and/or stomach distension. Intragastric administration of milk reproduced the decline in available receptors induced by suckling, whereas stomach distension produced by saline was ineffective. Concomitant with the decline in receptors, suckling produced a significant increase in both stomach content and plasma levels of an LHRH-like material. Fractionation of acid milk extracts in Sephadex G-25 yielded two peaks, one in a position similar to that of synthetic LHRH. Milk LHRH purified in a C-18 Sep-Pak column displaced binding of [125I]A-LHRH to ovarian membranes and yielded a parallel binding curve in the LHRH RIA. This milk LHRH also released LH and FSH from pituitaries in vitro and inhibited FSH-induced estradiol and progesterone secretion from granulosa cells in culture in a dose-related manner, comparable to A-LHRH. The suckling-induced decrease in available LHRH receptors was prevented by iv administration of a specific LHRH antiserum directed against all 10 amino acids of native LHRH, suggesting that milk LHRH reaches the ovary via the bloodstream. The results indicate that an LHRH-like substance of milk origin binds to LHRH receptors in the pup ovary, thereby controlling receptor availability.(ABSTRACT TRUNCATED AT 400 WORDS)

Essential fatty acid deficiency delays the onset of puberty in the female rat.

This study assessed the effect of a dietary deficiency in the essential fatty acids (EFA) linoleic and linolenic acids on the onset of female puberty. EFA deficiency was produced in female rats by means of a semipurified diet and was biochemically documented by analyzing serum and erythrocyte fatty acid levels of more than 30 fatty acids, including all members of the n-6 and n-3 series. Levels of linoleic acid (18:2 n-6) and all n-6 derivatives, particularly arachidonic acid, were strikingly reduced. A less pronounced but clear-cut decrease in n-3 fatty acids, including docosahexaenoic acid (22:6 n-3) was also found. The times of puberty and first ovulation, as assessed by the ages at vaginal opening and first diestrus, were significantly delayed in EFA-deficient rats. The mechanisms underlying this delay appear to reside at both hypothalamic and ovarian sites. Simulation of preovulatory plasma estradiol (E2) levels via implantation of E2-containing Silastic capsules evoked a LH surge 30 h later in control juvenile rats, but not in EFA-deficient animals, indicating a delay in the development of the hypothalamic component of E2-positive feedback in the latter group. This delay appears to be due at least in part to reduced prostaglandin E2 (PGE2) synthesis, as the ability of the neurotransmitter norepinephrine to induce PGE2 release from median eminence nerve terminals was markedly reduced in EFA-deficient rats compared with that in controls. The decrease in hypothalamic PGE2 release was related to the EFA deficiency and not to reduced PG synthase activity, as determined by HPLC analysis of PG synthase products derived from exogenous [14C]arachidonic acid. Basal and hCG-stimulated PGE2 synthesis was also compromised in ovaries from EFA-deficient rats. Depressed gonadal function resulting from the EFA deficiency was further evidenced by a reduced gonadotropin receptor content, a blunted E2 response to hCG in vitro, and an increase in mean serum FSH levels. These results suggest that the delay in puberty resulting from EFA deficiency is due to a reduced availability of arachidonic acid for synthesis of bioactive metabolites. This results in delayed development of both the hypothalamic and ovarian components of the reproductive axis.

The two-dimensional restriction analysis of Drosophila DNAs: males and females.

Two-dimensional restriction analysis has been applied to terminally labeled restriction segments from adult male and female flies that were the progeny of ten successive single-pair, brother-sister matings. Many of the 2500-4000 different bands that can be seen on autoradiographs are of widely different density, a fact that suggests the superposition of multiple restriction segments of similar sequence. The number of bands observed is significantly fewer than expected. Nearly all of the bands seen in male-DNA displays appear equivalent to those seen in female-DNA displays, but a few sex differences (male-dense and female-dense bands) can be identified--much fewer than expected. When the HpaII-MspI test was applied to fractionated BamHI fragments, between 900 and 1800 discrete C-C-G-G sites were found to be cleaved in a completely equivalent fashion by each isoschizomer.