Maternal cocaine administration in mice alters DNA methylation and gene expression in hippocampal neurons of neonatal and prepubertal offspring

Previous studies documented significant behavioral changes in the offspring of cocaine-exposed mothers. We now explore the hypothesis that maternal cocaine exposure could alter the fetal epigenetic machinery sufficiently to cause lasting neurochemical and functional changes in the offspring. Pregnant CD1 mice were administered either saline or 20 mg/kg cocaine twice daily on gestational days 8-19. Male pups from each of ten litters of the cocaine and control groups were analyzed at 3 (P3) or 30 (P30) days postnatum. Global DNA methylation, methylated DNA immunoprecipitation followed by CGI(2) microarray profiling and bisulfite sequencing, as well as quantitative real-time RT-PCR gene expression analysis, were evaluated in hippocampal pyramidal neurons excised by laser capture microdissection. Following maternal cocaine exposure, global DNA methylation was significantly decreased at P3 and increased at P30. Among the 492 CGIs whose methylation was significantly altered by cocaine at P3, 34% were hypermethylated while 66% were hypomethylated. Several of these CGIs contained promoter regions for genes implicated in crucial cellular functions. Endogenous expression of selected genes linked to the abnormally methylated CGIs was correspondingly decreased or increased by as much as 4-19-fold. By P30, some of the cocaine-associated effects at P3 endured, reversed to opposite directions, or disappeared. Further, additional sets of abnormally methylated targets emerged at P30 that were not observed at P3. Taken together, these observations indicate that maternal cocaine exposure during the second and third trimesters of gestation could produce potentially profound structural and functional modifications in the epigenomic programs of neonatal and prepubertal mice

Auditory Cortex Responses to Clicks and Sensory Modulation Difficulties in Children with Autism Spectrum Disorders (ASD)

Auditory sensory modulation difficulties are common in autism spectrum disorders (ASD) and may stem from a faulty arousal system that compromises the ability to regulate an optimal response. To study neurophysiological correlates of the sensory modulation difficulties, we recorded magnetic field responses to clicks in 14 ASD and 15 typically developing (TD) children. We further analyzed the P100m, which is the most prominent component of the auditory magnetic field response in children and may reflect preattentive arousal processes. The P100m was rightward lateralized in the TD, but not in the ASD children, who showed a tendency toward P100m reduction in the right hemisphere (RH). The atypical P100m lateralization in the ASD subjects was associated with greater severity of sensory abnormalities assessed by Short Sensory Profile, as well as with auditory hypersensitivity during the first two years of life. The absence of right-hemispheric predominance of the P100m and a tendency for its right-hemispheric reduction in the ASD children suggests disturbance of the RH ascending reticular brainstem pathways and/or their thalamic and cortical projections, which in turn may contribute to abnormal arousal and attention. The correlation of sensory abnormalities with atypical, more leftward, P100m lateralization suggests that reduced preattentive processing in the right hemisphere and/or its shift to the left hemisphere may contribute to abnormal sensory behavior in ASD

Neonatal local noxious insult affects gene expression in the spinal dorsal horn of adult rats

Neonatal noxious insult produces a long-term effect on pain processing in adults. Rats subjected to carrageenan (CAR) injection in one hindpaw within the sensitive period develop bilateral hypoalgesia as adults. In the same rats, inflammation of the hindpaw, which was the site of the neonatal injury, induces a localized enhanced hyperalgesia limited to this paw. To gain an insight into the long-term molecular changes involved in the above-described long-term nociceptive effects of neonatal noxious insult at the spinal level, we performed DNA microarray analysis (using microarrays containing oligo-probes for 205 genes encoding receptors and transporters for glutamate, GABA, and amine neurotransmitters, precursors and receptors for neuropeptides, and neurotrophins, cytokines and their receptors) to compare gene expression profiles in the lumbar spinal dorsal horn (LDH) of adult (P60) male rats that received neonatal CAR treatment within (at postnatal day 3; P3) and outside (at postnatal 12; P12) of the sensitive period. The data were obtained both without inflammation (at baseline) and during complete Freund's adjuvant induced inflammation of the neonatally injured paw. The observed changes were verified by real-time RT-PCR. This study revealed significant basal and inflammation-associated aberrations in the expression of multiple genes in the LDH of adult animals receiving CAR injection at P3 as compared to their expression levels in the LDH of animals receiving either no injections or CAR injection at P12. In particular, at baseline, twelve genes (representing GABA, serotonin, adenosine, neuropeptide Y, cholecystokinin, opioid, tachykinin and interleukin systems) were up-regulated in the bilateral LDH of the former animals. The baseline condition in these animals was also characterized by up-regulation of seven genes (encoding members of GABA, cholecystokinin, histamine, serotonin, and neurotensin systems) in the LDH ipsilateral to the neonatally-injured paw. The largest aberration in gene expression, however, was observed during inflammation of the neonatally injured hindpaws in the ipsilateral LDH, which included thirty-six genes (encoding numerous members of glutamate, serotonin, GABA, calcitonin gene-related peptide, neurotrophin, and interleukin systems). These findings suggest that changes in gene expression may be involved in the long-term nociceptive effects of neonatal noxious insult at the spinal level

Thidiazuron-induced formation of strawberry microshoots on different nutrient media

The present study was aimed to improve in vitro shoot proliferation of octoploid strawberry cultivar ‘Solnechnaya polyanka’ on different nutrient media (MS, B5 or MS+B5), supplemented with various concentrations (2.0, 4.0, 8.0 ȝM) of thidiazuron (TDZ). The best regeneration frequency (100%) with the highest rate of strawberry axillary shoot proliferation (14.1 ± 1.24 shoots per explant) and maximum shoot length (1.92 ± 0.04 cm) were achieved using precultivation of explants for 3 days on induction media B5 supplemented with 4.0 ȝM TDZ followed by cultivation of regenerants on hormone-free media B5 for 6 weeks

How Can the Engineering Parameters of the NIR Grader Affect the Efficiency of Seed Grading?

The automated grading of Scots pine seeds in the near-infrared wavelength region (NIR grading) is a starting point for further actions, such as coating and priming. This reduces the time and financial costs and increases the accuracy of seed viability classification compared to invasive techniques. The NIR-based wave reflected from each pine seed must be detected and processed with sufficient accuracy. To focus the reflected beam, we used fiber-optic Bragg grating, a Bragg mirror, and diffraction grating. For each focusing option based on the DOE matrix, one experiment of 20 runs (n = 20) and three replicas (m = 3) in each run was conducted. In each replica, we used 100 conditioned and 100 non-conditioned seeds (NC + NNC = 200) selected randomly from five samples weighing 50 g from a seedlot weighing 1 kg extracted from cones collected from a natural tree stand. Three experiments were conducted on the NIR grading of Scots pine seeds using an optoelectronic device. An adequate DOE regression model of the grading efficiency function was obtained. The functions included the following arguments: angle of incidence of the optical beam, NIR wavelength reflected from the seed, and height of the seed pipeline. The influence of the inclination angle of the light source relative to the plane of pine seed movement on the grading quality prevails over other factors. The NIR grading of Scots pine seeds allows the separation of seeds according to the viability index, which is important, since dead petrified seeds (possibly up to 25%) may occur in the seed batch, which cannot be eliminated by either seed size or mass. The peak of NIR grading is achieved by combining the average grader engineering parameters: 968–973 nm for the wavelength and 44–46 degrees for the inclination angle of the reflected beam at a seed pipe size of 0.18–0.23 m

An innovative approach to ex vitro rooting and acclimatization of Fragaria × ananassa Duch. microshoots using а biogenic silica- and green-tea-catechin-based mechanocomposite

A new approach for rapid ex vitro rooting and acclimatization of Fragaria × ananassa micropropagated plantlets of two cultivars (“Alpha” and “Festivalnaya”) has been developed using a mechanocomposite based on biogenic silica and green-tea catechins. Two different mechanocomposite treatments were studied: dipping the cut ends of microshoots in the mechanocomposite powder (the dry dip method) and single watering with solutions at concentrations of 0.3, 1.0, and 3.0 g L⁻¹. These variants were compared with pulse treatment of microplants with 30 mg L⁻¹ indole-3-acetic acid (IAA) for 4 h and a control group of microshoots that were moistened with hormone-free ¼-strength MS medium. The frequencies of ex vitro rooting at the end of the acclimatization period (30 d) varied from 24.8 to 99.7%. The dry dip treatment was best (rooting frequency about 100%) with up to 7.15 ± 0.54-cm root length, and 6.10 ± 0.31 roots per plantlet. Moreover, this study showed that the growth-stimulating effect of this mechanocomposite treatment on root formation resulted in increased rosette height, leaf number, leaf area, and dry weight of aerial parts. Histological analysis of the leaf blades revealed decreased mesophyll thickness of microshoots treated with the mechanocomposite (up to 88.77 ± 2.95 vs. 111.51 ± 3.56 μm for the control). Morphometric analysis of scanning electron microscopy data showed that mechanocomposite treatments led to increased stomata density and stomata length. These structural changes led to normalization of the water regime and indicated successful acclimatization. The combination of ex vitro rooting and acclimatization reduced the procedure time by 4 wk, and may be used for commercial strawberry micropropagation

Wood Quality along the Trunk Height of Birch and Aspen Growing in the Restoring Forests of Central Russia

The structure of forests has changed with an increase in soft-wooded broadleaved species over the past decade. The demand for hard-wooded broadleaved species can be met by replacing them with compressed wood of soft-wooded broadleaved species. Existing compressed wood technologies do not fully take into account the density variations that exist along the height of a tree trunk. In this study, we examined the variability of birch and aspen microstructures along the height of the trunk, including vessels per square millimeter and the diameter (tangential and radial) of the vessel lumina. The research was carried out on aspen and birch species growing in Central Russia. The vessels per square millimeter in both species increased from the base to the top of the trunk and their diameters decreased from the base to the top of the trunk. Birch demonstrated greater changes in these values than aspen. There was a strong relationship between the diameter of the vessel lumina and the trunk height. A decrease in the density of the stemwood from the base to the top of the trunk was caused by an increase in the vessels per square millimeter. These results affected the density of the stemwood and determined the degree of compression as well as the initial size of the blanks required to obtain material with uniform quality indicators, regardless of the source location of the raw materials in the tree trunk

Effects of maternal cocaine exposure on CGI DNA methylation in P30 compared to P3 pups.

<p>(a) Cocaine-induced hypermethylated and hypomethylated CGIs in P30 compared to P3 pups. Shown are the numbers of CGIs with altered methylation levels in cocaine-treated animals relative to saline-controls at each age. For P30 data, the number of CGIs with altered methylation levels that did not show a change at P3 (<i>new</i>) are indicated by a cross-hatched segment of the bar. Also indicated by differential shading are CGIs that were significantly altered at both P3 and P30, retaining or reversing their direction of change at P30 compared to P3 (<i>retained direction; changed direction</i>). (b) Hypermethylated and hypomethylated promoter-associated CGIs at P3 and P30. For both ages, CGIs associated with promoters of neural tissue-specific genes (<i>neural genes</i>) are represented by white bars, while CGIs associated with promoters of non-neural tissue-specific genes (<i>housekeeping genes</i>) are represented by black bars. In addition, for P30, the number of CGIs with altered methylation state undetectable at P3 (<i>new</i>) and the number of CGIs corresponding to the CGIs affected at P3, which either retained or reversed the direction of change (<i>changed direction/retained direction</i>), are indicated as in a. (c) Abnormally methylated repetitive element-associated CGIs at P3 and P30.</p

CpG methylograms of selected abnormally methylated promoters in P3 mice.

<p>From the MeDIP/CGI array-generated methylation profiles, ten cocaine-induced abnormally methylated CGIs known to be associated with gene promoter regions were selected and cloned, followed by bisulfite sequencing to identify the relative locations of methylated CpGs in the gene promoter sequence. For each chart, the name of the promoter is shown on the left vertical while the horizontal numbers mark the CpGs on the sequence; the CpGs are numbered from the 5'end of the clone. The vertical numbers on the right (1–5) indicate data rows for samples from each of 5 animals from the control (upper, DRN) and cocaine (lower, COC) groups. For each sample, three clones were sequenced, hence the three sub-rows available to each sample for methylation mapping. For each sub-row representing a clone, white squares indicate unmethylated CpGs while black-filled squares indicate methylated CpGs. If all three clones were consistent, the filled boxes would appear for each sub-row for that sample. If all 5 samples (mice) were consistent, then the filled boxes would extend vertically across all 5 rows. The relative density of filled boxes between the control (upper 5) samples and the cocaine (lower 5) samples provides a visual feedback on the relative levels of CpG methylation in these DNA regions. For example, clones of DYRK3 or COQ7 promoters contained lower proportions of methylated CpGs in the cocaine-treatment pups than in the saline-control offspring, whereas the converse applies in the case of GPR73 or MAPK1 clones.</p