The effects of vitamin D3 during pregnancy and lactation on offspring physiology and behavior in sprague-dawley rats.

Recent findings show that developmental vitamin D deficiency leads to altered brain morphology and behavioral development in the rat offspring. We examined the effects of different dietary vitamin D levels in rat dams on behavior and biochemistry of the offspring. Females were divided into five conditions and received diets containing 0, 1,5, 3.3, 6.0, or 10.0 IU/g of vitamin D3 from mating to weaning. Offspring were tested as juveniles and as adults for anxiety, social learning and behavior, and locomotion. Results show that both deficient and excessive levels of vitamin D3 in juveniles lead to altered physiology and behavior. In juveniles but not adults, variations in vitamin D were related to variations in measures of anxiety and marginally, activity levels. For social behaviors, both juveniles and adults were affected by mothers' diets. In general, offspring of animals receiving abnormal concentrations of vitamin D showed the most deficits.

Dendritic morphology in the striatum and hypothalamus differentially exhibits experience-dependent changes in response to maternal care and early social isolation.

Changes in neuron morphology, stemming from experiences in early life or adulthood, may be the basis for changes in behavior and their underlying functional mechanisms. For example, reproductive experience has been shown to significantly alter neuron morphology in the hippocampus and prefrontal cortex. In contrast to the effects of reproductive experience, a form of enrichment, on neuron morphology, our understanding of the effects of early social isolation on adult neuron morphology is limited. Therefore, the present study examined changes in neuron morphology in the dorsal (caudate nucleus) and ventral (nucleus accumbens, shell region) striatum and the medial preoptic area of adult virgin and postpartum females exposed to either artificial or maternal rearing during development. Primary results show that regardless of early social isolation, neurons in the caudate nucleus of postpartum females have decreased dendritic complexity compared to virgin females. Maternal experience also increased dendritic complexity in neurons of the nucleus accumbens shell. However, both early social isolation and maternal experience in adulthood influenced dendritic complexity in the medial preoptic area. Together these findings suggest that hypothalamic and striatal neurons show experience-dependent dendritic plasticity and the type and timing of these experiences differentially affect the location and degree of these morphological changes.

Artificial rearing inhibits apoptotic cell death through action on pro-apoptotic signaling molecules during brain development: replacement licking partially reverses these effects.

Early life stress associated with being reared without mother, siblings, and nest affects the formation of neuronal networks during rat development. Prior work shows that in comparison to mother-reared male rats, artificial rearing results in elevated numbers of neurons in adulthood and reduced apoptosis during the first postnatal week. Replacement with stroking stimulation, designed to simulate mothers' licking, reversed these effects in most brain areas. The present communication explored the effects of early rearing manipulations on signaling proteins. Male rats were reared until postnatal day 7 either in an artificial-feeding paradigm (AR) or with their mothers (MR). AR animals received different amounts of maternal-like stimulation using a soft paintbrush. Brains were extracted and prepared for molecular assays of 1) apoptosis and 2) pro and anti-apoptotic proteins on day 7 of postnatal life. Results showed that stimulation of the AR pups reversed the effects of artificial rearing on apoptosis in a dose dependent manner; low and very high levels of stimulation were without effect whereas moderate levels of stimulation produced effects on apoptosis similar to effects seen in mother-reared controls. Moreover, this artificial rearing effect and the pattern of reversal with stroking were also found for levels of pro-apoptotic Bax protein, the ratio of Bax/Bcl-2 and levels of activated caspase-3 which we believe mediates programmed cell death.

Prenatal restraint stress and motherless rearing disrupts expression of plasticity markers and stress-induced corticosterone release in adult female Sprague-Dawley rats.

This study investigated the effects of prenatal stress and complete maternal deprivation, using the artificial rearing (AR) paradigm, on the expression of neural plasticity markers and hypothalamic-pituitary-adrenal (HPA) axis responsivity to stress. Rats were exposed to stress during gestation (day 10-21) and postnatally were either artificially reared (AR) or mother reared (MR). AR involves complete separation of the pup from both the dam and the litter throughout the pre-weaning period. In adulthood, we measured levels of corticosterone (CORT) in response to restraint stress. Also, we examined the expression of synaptophysin (SYN) and brain-derived neurotrophic factor (BDNF) in the medial prefrontal cortex (MPFC) and the nucleus accumbens (Nacc), areas of the brain that mediate behavioral activation and attention, among other behaviors. Earlier work on the same rats indicated that these behavioral endpoints, such as locomotor activity and sensorimotor gating, are affected by our prenatal and postnatal manipulations. Prenatal stress decreased CORT at 20 and 90 min post-stressor in MR, but not in AR, animals. Also, in comparison to MR groups, AR decreased SYN and BDNF expression in the MPFC and Nacc. Additional somatosensory 'licking-like' stroking stimulation partially reversed the effects of AR. Prenatal stress did not have a robust main effect but affected the impact of the postnatal rearing condition on SYN expression and stress-induced CORT. These results suggest that both prenatal and postnatal adversities have an influence on HPA axis responsivity and alter the expression of plasticity related neuronal proteins.

Maternal isolation alters the expression of neural proteins during development: 'Stroking' stimulation reverses these effects.

Rat pups reared apart from their siblings, mother, and nest environment in the 'pup-in-a-cup' regime show many alterations in behavior reminiscent of the Institutional Inattention/Overactivity Syndrome that characterizes children whose first few months are spent in institutions. In this report, we compare mother-reared (MR) and artificially reared (AR) male rats in concentrations and distributions of brain proteins that are involved in normal brain development. When assessed during the juvenile period and in adulthood, AR animals showed elevations in Neu-N (a neuronal marker) and in S-100 (an astrocyte marker) but reductions in synaptophysin (synapse protein), N-CAM (cell-adhesion molecule), GAP-43 (axon elongation protein), and BDNF (brain derived neurotrophic factor) in comparison to MR controls in many brain sites involved in attention, impulsivity, activity, and social behavior. Daily 'licking-like' stimulation provided to AR animals (AR-MAX) throughout early development that reverses many of the behavioral deficits, also reverses many of the isolation effects on brain proteins. Study 2 showed that elevations in the number of neurons in combination with decreases in functionality are associated with a reduction in neuronal pruning and apoptosis during the very early post-partum period in AR animals and their reversal through daily 'licking-like' stimulation.

Isolation, characterization, and bioinformatic analysis of calmodulin-binding protein cmbB reveals a novel tandem IP22 repeat common to many Dictyostelium and Mimivirus proteins.

A novel calmodulin-binding protein cmbB from Dictyostelium discoideum is encoded in a single gene. Northern analysis reveals two cmbB transcripts first detectable at 4 h during multicellular development. Western blotting detects an approximately 46.6 kDa protein. Sequence analysis and calmodulin-agarose binding studies identified a "classic" calcium-dependent calmodulin-binding domain (179IPKSLRSLFLGKGYNQPLEF198) but structural analyses suggest binding may not involve classic alpha-helical calmodulin-binding. The cmbB protein is comprised of tandem repeats of a newly identified IP22 motif ([I,L]Pxxhxxhxhxxxhxxxhxxxx; where h = any hydrophobic amino acid) that is highly conserved and a more precise representation of the FNIP repeat. At least eight Acanthamoeba polyphaga Mimivirus proteins and over 100 Dictyostelium proteins contain tandem arrays of the IP22 motif and its variants. cmbB also shares structural homology to YopM, from the plague bacterium Yersenia pestis.

Isolation and characterization of Dictyostelium thymidine kinase 1 as a calmodulin-binding protein.

Probing of a cDNA expression library from multicellular development of Dictyostelium discoideum using a recombinant radiolabelled calmodulin probe (35S-VU1-CaM) led to the isolation of a cDNA encoding a putative CaM-binding protein (CaMBP). The cDNA contained an open reading frame of 951 bp encoding a 227aa polypeptide (25.5 kDa). Sequence comparisons led to highly significant matches with cytosolic thymidine kinases (TK1; EC 2.7.1.21) from a diverse number of species including humans (7e-56; 59% Identities; 75% Positives) indicating that the encoded protein is D. discoideum TK1 (DdTK1; ThyB). DdTK1 has not been previously characterized in this organism. In keeping with its sequence similarity with DdTK1, antibodies against humanTK1 recognize DdTK1, which is expressed during growth but decreases in amount after starvation. A CaM-binding domain (CaMBD; 20GKTTELIRRIKRFNFANKKC30) was identified and wild type DdTK1 plus two constructs (DdTK deltaC36, DdTK deltaC75) possessing the domain were shown to bind CaM in vitro but only in the presence of calcium while a construct (DdTK deltaN72) lacking the region failed to bind to CaM. Thus, DdTK1 is a Ca2+-dependent CaMBP. Sequence alignments against TK1 from vertebrates to viruses show that CaM-binding region is highly conserved. The identified CaMBD overlaps the ATP-binding (P-loop) domain suggesting CaM might affect the activity of this kinase. Recombinant DdTK is enzymatically active and showed stimulation by CaM (113+/-0.5%) an in vitro enhancement that was prevented by co-addition of the CaM antagonists W7 (91.2+/-0.8%) and W13 (96.6+/-0.6%). The discovery that TK1 from D. discoideum, and possibly other species including humans and a large number of human viruses, is a Ca2+-dependent CaMBP opens up new avenues for research on this medically relevant protein.