Indirect Effects of Glucagon-Like Peptide-1 Receptor Agonist Exendin-4 on the Peripheral Circadian Clocks in Mice

Circadian clocks in peripheral tissues are powerfully entrained by feeding. The mechanisms underlying this food entrainment remain unclear, although various humoral and neural factors have been reported to affect peripheral clocks. Because glucagon-like peptide-1 (GLP-1), which is rapidly secreted in response to food ingestion, influences multiple humoral and neural signaling pathways, we suggest that GLP-1 plays a role in the food entrainment of peripheral clocks. To test this, we compared the effects of exendin-4, a GLP-1 receptor agonist, on mRNA expression of the clock genes (Clock, Bmal1, Nr1d1, Per1, Per2, and Cry1) with those of refeeding. In addition, we investigated whether exendin-4 could affect the rhythms of the peripheral clocks. In male C57BL/6J mice, although refeeding rapidly (within 2 h) altered mRNA levels of Per1 and Per2 in the liver and that of Per1 in adipose tissue, a single i.p. injection of exendin-4 did not cause such changes. However, unlike the GLP-1 receptor antagonist exendin-(9–39), exendin-4 significantly influenced Per1 mRNA levels in the liver at 12 h after injection. Moreover, pretreatment with exendin-4 affected the rapid-feeding-induced change in Per1 not only in the liver, but also in adipose tissue, without effect on food intake. Furthermore, during light-phase restricted feeding, repeated dosing of exendin-4 at the beginning of the dark phase profoundly influenced both the food intake and daily rhythms of clock gene expression in peripheral tissues. Thus, these results suggest that exendin-4 modulates peripheral clocks via multiple mechanisms different from those of refeeding.     

CCL-1 in the spinal cord contributes to neuropathic pain induced by nerve injury

Cytokines such as interleukins are known to be involved in the development of neuropathic pain through activation of neuroglia. However, the role of chemokine (C-C motif) ligand 1 (CCL-1), a well-characterized chemokine secreted by activated T cells, in the nociceptive transmission remains unclear. We found that CCL-1 was upregulated in the spinal dorsal horn after partial sciatic nerve ligation. Therefore, we examined actions of recombinant CCL-1 on behavioural pain score, synaptic transmission, glial cell function and cytokine production in the spinal dorsal horn. Here we show that CCL-1 is one of the key mediators involved in the development of neuropathic pain. Expression of CCL-1 mRNA was mainly detected in the ipsilateral dorsal root ganglion, and the expression of specific CCL-1 receptor CCR-8 was upregulated in the superficial dorsal horn. Increased expression of CCR-8 was observed not only in neurons but also in microglia and astrocytes in the ipsilateral side. Recombinant CCL-1 injected intrathecally (i.t.) to naive mice induced allodynia, which was prevented by the supplemental addition of N-methyl-𝒟-aspartate (NMDA) receptor antagonist, MK-801. Patch-clamp recordings from spinal cord slices revealed that application of CCL-1 transiently enhanced excitatory synaptic transmission in the substantia gelatinosa (lamina II). In the long term, i.t. injection of CCL-1 induced phosphorylation of NMDA receptor subunit, NR1 and NR2B, in the spinal cord. Injection of CCL-1 also upregulated mRNA level of glial cell markers and proinflammatory cytokines (IL-1β, TNF-α and IL-6). The tactile allodynia induced by nerve ligation was attenuated by prophylactic and chronic administration of neutralizing antibody against CCL-1 and by knocking down of CCR-8. Our results indicate that CCL-1 is one of the key molecules in pathogenesis, and CCL-1/CCR-8 signaling system can be a potential target for drug development in the treatment for neuropathic pain.     

Evidence of intragenic recombination in G1 rotavirus VP7 genes

The G1 rotavirus is the most widespread genotype causing acute gastroenteritis in children. In an attempt to investigate the occurrence of intragenic recombination, 131 complete coding region sequences of VP7 genes of the G1 rotaviruses in GenBank were examined. Three hitherto-unreported intragenic recombinant rotaviruses were identified. It was noteworthy that two different types (interlineage and intersublineage) of intragenic recombination in rotaviruses were also found. This is the first report to demonstrate the existence of intragenic recombinations between interlineage and intersublineage in G1 rotaviruses.     

Decrease in the Traumatic Symptoms Observed in Child Survivors within Three Years of the 2011 Japan Earthquake and Tsunami

Background

On March 11, 2011, Japan was struck by a massive earthquake and tsunami. The tsunami caused tremendous damage and traumatized several people, including children. The aim of this study was to assess changes in traumatic symptoms 8, 20, and 30 months of the 2011 tsunami.

Methods

The study comprised three groups. Copies of the Post-Traumatic Stress Symptoms for Children 15 items (PTSSC-15), a self-rating questionnaire on traumatic symptoms, were distributed to 12,524 children (8-month period), 12,193 children (20-month period), and 11,819 children (30-month period). An effective response of children 8 months, 20 months, and 30 month after the disaster was obtained in 11,639 (92.9%), 10,597 (86.9%), and 10,812 children (91.4%), respectively. We calculated the total score, PTSD subscale, and Depression subscale of PTSSC-15. We calculated the total score, PTSD subscale, and Depression subscale of PTSSC-15.

Results

The PTSSC-15 total score and PTSD subscale of children belonging to 1st–9th grade groups who were tested 30 and 20 months after the tsunami significantly decreased compared with those of children tested 8 months after the tsunami. The PTSSC-15 total score and PTSD subscale of children in 1st–9th grade groups tested after 30 months did not decrease significantly compared with those of children tested after 20 months. The PTSSC-15 Depression subscale and PTSD subscale of children in 1st–9th grade groups tested after 30 months significantly decreased compared with those of children tested 8 months after the tsunami. The PTSSC-15 Depression subscale of children in 1st–9th grade groups evaluated after 30 months significantly decreased compared with those of children evaluated after 20 months.

Conclusions

This study demonstrates that the traumatic symptoms of children who survived the massive tsunami improved with time. Nonetheless, the traumatic symptoms, which in some cases did not improve with time.     

Bovine adrenocortical adenylate cyclase: Some properties of the solubilized,fluoride-activated enzyme

The activity of bovine adrenocortical plasma membrane adenylate cyclase can be maintained at 4°C in the presence of NaF. The half-life of the fluoride-stabilized enzyme is approximately 7 days. Maximal activation by fluoride requires approximately 20 min at 0°C and the level of activity attained is dependent on fluoride concentration. The enzyme from freshly harvested membranes can also be stimulated by ACTH_{1 – 24} and Gpp(NH)p and the stimulatory effects of these two activators are additive. Prolonged exposure to either NaF or Gpp(NH)p precludes hormone activation. Optimal concentration for Gpp(NH)p activation is 10^?4–10^?5M. Treatment of the enzyme with a Tris-HCl buffer containing Lubrol-PX (1%), NaF, dithiothreitol, and MgSO₄ followed by sonication affords a preparation that does not sediment at 100 000 g in 1 hr. This material has a low specific activity; however, removal of the detergent on DEAE cellulose restores specific activity to its original level. A significant improvement in specific activity is observed following dialysis or ultrafiltration of the detergent-free 100 000-g supernatant. At this stage the enzyme can be lyophilized and stored at ?70°C without loss of activity. The enzyme in the detergent-free 100 000-g supernatant behaves as a single peak that is included in Sepharose 6B. Comparison of the elution profile of the enzyme with profiles produced by a standard set of proteins suggests a molecular weight of 1 × 10⁶. Hydrophobic chromatography of the detergent-free 100 000-g supernatant on n-hexyl Sepharose 4B results in a fivefold enhancement of specific activity.