Long-term depression-inductive stimulation causes long-term potentiation in mouse Purkinje cells with a mutant thyroid hormone receptor.

Thyroid hormones (THs) regulate gene expression by binding to nuclear TH receptors (TRs) in the cell. THs are indispensable for brain development. However, we have little knowledge about how congenital hypothyroidism in neurons affects functions of the central nervous system in adulthood. Here, we report specific TH effects on functional development of the cerebellum by using transgenic mice overexpressing a dominant-negative TR (Mf-1) specifically in cerebellar Purkinje cells (PCs). Adult Mf-1 mice displayed impairments in motor coordination and motor learning. Surprisingly, long-term depression (LTD)-inductive stimulation caused long-term potentiation (LTP) at parallel fiber (PF)-PC synapses in adult Mf-1 mice, although there was no abnormality in morphology or basal properties of PF-PC synapses. The LTP phenotype was turned to LTD in Mf-1 mice when the inductive stimulation was applied in an extracellular high-Ca2+ condition. Confocal calcium imaging revealed that dendritic Ca2+ elevation evoked by LTD-inductive stimulation is significantly reduced in Mf-1 PCs but not by PC depolarization only. Single PC messenger RNA quantitative analysis showed reduced expression of SERCA2 and IP3 receptor type 1 in Mf-1 PCs, which are essential for mGluR1-mediated internal calcium release from endoplasmic reticulum in cerebellar PCs. These abnormal changes were not observed in adult-onset PC-specific TH deficiency mice created by adeno-associated virus vectors. Thus, we propose the importance of TH action during neural development in establishing proper cerebellar function in adulthood, independent of its morphology. The present study gives insight into the cellular and molecular mechanisms underlying congenital hypothyroidism-induced dysfunctions of central nervous system and cerebellum.

The neurotoxic effect of lactational PFOS exposure on cerebellar functional development in male mice.

Recent studies showed a possible association between perfluorooctane sulfonate (PFOS) and developmental disabilities. We previously found the specific effects of PFOS exposure on learning and memory, however, its effect on the other developmental disabilities such as motor and social deficits remains unclear. We examined the effect of early lactational PFOS exposure on motor coordination, social activity, and anxiety in male mice. We orally administered a PFOS solution to dams from postnatal day 1-14. At 10 weeks old, we conducted a behavior test battery to evaluate motor performance, social activity, and anxiety, followed by electrophysiology and Western blot analysis. PFOS-exposed mice displayed impaired motor coordination. Whole-cell patch-clamp recordings from Purkinje cells revealed that the short-term and long-term plasticity at parallel fiber-Purkinje cell synapses are affected by PFOS exposure. Western blot analysis indicated that PFOS exposure increased syntaxin binding protein 1 (Munc18-1) and glutamate metabotropic receptor 1 (mGluR1) protein levels, which may be associated with the change in neurotransmitter release from parallel fibers and the level of long-term depression, respectively. The present study demonstrates that lactational PFOS exposure may have disrupted the pre- and postsynaptic plasticity at parallel fiber-Purkinje cell synapses, causing profound, long-lasting abnormal effects on the cerebellar function.

Neurotoxic effects of lactational exposure to perfluorooctane sulfonate on learning and memory in adult male mouse.

The present study aims to examine the effect of early lactational perfluorooctane sulfonate (PFOS) exposures on learning and memory in male mice and reveal the underlying mechanisms involved. PFOS solution was orally administered to dams from the postpartum days 1-14, so that pups would be exposed through breast milk. At 8-10 weeks of age, we performed object location test (OLT), object recognition test (ORT), and pairwise visual discrimination (VD) task. We also performed in vivo microdialysis, and mRNA and protein analysis of genes responsible for hippocampal development and function. In both OLT and ORT, the performance of mice in the PFOS-exposed group was significantly lower than those in the control group. In the VD task, the PFOS-exposed group learned significantly slower than the control group. Concentrations of glutamate and gamma-aminobutyric acid in the dorsal hippocampus were significantly higher in the PFOS-exposed group than in the control group. No notable differences were shown in our mRNA and protein studies. The present study showed that lactational PFOS exposure has a profound, long-lasting neurotoxic effect in the hippocampus and consequently leads to learning and memory deficits.

In Utero and Postnatal Propylthiouracil-Induced Mild Hypothyroidism Impairs Maternal Behavior in Mice.

Thyroid hormones (THs) play crucial roles in general and brain development. Even if the hypothyroidism is mild, it may alter brain function, resulting in irreversible behavioral alterations. Although various behavioral analyses have been conducted, the effects of propylthiouracil (PTU) treatment during in utero and postnatal periods on maternal behavior have not yet been studied. The present study examined in mice whether THs insufficiency during development induce behavioral changes. Pregnant C57BL/6j mice were divided into three groups, and each group was administered different dosages of PTU (0, 5, or 50 ppm) in drinking water during in utero and postnatal periods (from gestational day 14 to postnatal day 21). First, locomotor activity and cognitive function were assessed in the offspring at 10 weeks. Next, female offspring were mated with normal mice and they and their offspring were used to assess several aspects of maternal behavior (identifying first pup, returning all pups to nest, time spent nursing, and licking pups). As expected, locomotor and cognitive functions in these mice were disrupted in a PTU dose-dependent manner. On postpartum day 2, dams who had been exposed 50 ppm PTU during in utero and postnatal periods displayed a significantly longer time identifying the first pup and returning all three pups back to the nest, less time nursing, and decreased licking behavior. The decrease in maternal behavior was significantly correlated with a decrease in cognition. These results indicate that insufficiency of THs during in utero and postnatal periods impairs maternal behavior, which may be partly induced by impaired cognitive function.

Melanoma antigen family A4 protein produced by transgenic silkworms induces antitumor immune responses.

Recent clinical trials with the aim of developing tumor antigen (TA)-specific cancer vaccines against a number of malignancies have focused on the identification of TAs presented by tumor cells and recognized by T cells. In the present study, the TA melanoma antigen family A4 (MAGE-A4) protein was produced using a transgenic (TG) silkworm system. Using in vitro stimulation, it was subsequently determined whether MAGE-A4 protein induced MAGE-A4-specific T cells from peripheral blood mononuclear cells of healthy donors. TG silkworm lines expressing a MAGE-A4 gene under an upstream activating sequence (UAS) were mated with those expressing a yeast transcription activator protein (GAL4) at the middle silk glands (MSGs) and embryos that harbored both the GAL4 and UAS constructs were selected. Recombinant MAGE-A4 protein was extracted from the MSGs of TG silkworms and evaluated using SDS-PAGE and western blot analysis. It was observed that MAGE-A4 produced by the TG silkworm system successfully induced MAGE-A4-specific CD4+ T cell responses. Furthermore, MAGE-A4-specific CD4+ T cells recognized antigen-presenting cells when pulsed with a MAGE-A4+ tumor cell lysate. The present data suggests that recombinant tumor antigen production using the TG silkworm system may be a novel tool in the preparation of cancer vaccines.

Effects of Mild Perinatal Hypothyroidism on Cognitive Function of Adult Male Offspring.

Mild perinatal hypothyroidism may result from inadequate iodine intake, insufficient treatment of congenital hypothyroidism, or exposure to endocrine-disrupting chemicals. Because thyroid hormones are critical for brain development, severe hypothyroidism that is untreated in infancy causes irreversible cretinism. Milder hypothyroidism may also affect cognitive development; however, the effects of mild and/or moderate hypothyroidism on brain development are not fully understood. In this study, we examined the behavior of adult male mice rendered mildly hypothyroid during the perinatal period using low-dose propylthiouracil (PTU). PTU was administered through drinking water (5 or 50 ppm) from gestational day 14 to postnatal day 21. Cognitive performance, studied by an object in-location test (OLT), was impaired in PTU-treated mice at postnatal week 8. These results suggest that, although the hypothyroidism was mild, it partially impaired cognitive function. We next measured the concentration of neurotransmitters (glutamate, γ-aminobutyric acid, and glycine) in the hippocampus using in vivo microdialysis during OLT. The concentrations of neurotransmitters, particularly glutamate and glycine, decreased in PTU-treated mice. The expression levels of N-methyl-d-aspartate receptor subunits, which are profound regulators of glutamate neurotransmission and memory function, also were decreased in PTU-treated mice. These data indicate that mild perinatal hypothyroidism causes cognitive disorders in adult offspring. Such disorders may be partially induced secondary to decreased concentrations of neurotransmitters and receptor expression.

Early-life stress induces motor coordination dysfunction in adult mice.

Although child abuse has become a serious social problem in most countries, the neural mechanisms by which it induces adulthood mental disorders is not yet fully understood. Mice exposed to early-life stresses, such as maternal deprivation (MD) during lactation, are a good model for studying the effects of neglect of humans in early life. Early-life stress induces structural/functional changes of neurons in the hippocampus, prefrontal cortex, and amygdala, and causes mental disorders in adulthood. In this study, we found motor coordination dysfunction in male MD mice. We also found that the expression levels of the aminomethylphosphonic acid receptor subunits GluA1 and GluA3 were high in the cerebellum of male MD mice. The basal activity of the cerebellum detected by field-potential analysis was higher in male MD mice than in male control mice. Caloric stimulation increased the activity of the cerebellum of control mice, but it did not significantly increase the activity of the cerebellum in male MD mice. We concluded that early-life stress induced a functional change in the cerebellum of MD mice and that this change induced motor coordination dysfunctions.

Early-life-stress affects the homeostasis of glutamatergic synapses.

Early-life stress induces several neuropsychological disorders in adulthood, including depression. Such disorders may be induced by functional alteration of the glutamatergic system. However, their underlying mechanisms have not yet been fully clarified. Furthermore, the involvement of glucocorticoids, which are representative stress hormones, has not yet been fully clarified. In this study, we used maternal deprivation (MD) mice as an early-life-stress model, and studied the changes in the glutamatergic system in adulthood. The glutamate concentration and neuronal activity in the somatosensory cortex (SSC) increased under basal conditions in MD mice. Stressful physical stimulation (SPS) increased the concentration of corticosterone, but not of glutamate, in the control mouse SSC. On the other hand, in the MD mice, although the basal concentration of corticosterone in the SSC increased, no SPS-induced increase was observed. In contrast, the concentration of glutamate increased greatly during SPS. It was significantly high for 30 min after stimulation. The expression level of α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid/N-methyl-d-aspartate receptors in the MD mice was also changed compared with that in the control mice after stimulation. These findings indicate that early-life stress disrupts the homeostasis of glutamatergic synapses.