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Activating transcription factor 6 (ATF6) is an endoplasmic reticulum (ER) stress-regulated transcription factor that induces expression of major molecular chaperones in the ER. We recently reported that ATF6β, a subtype of the ATF6, induced expression of calreticulin, a molecular chaperone in the ER with a high Ca2+-binding capacity, and promoted neuronal survival against ER stress and excitotoxicity. In the present study, we demonstrate that ATF6β deficiency in mice also decreases calreticulin expression and increases expression of glucose-regulated protein 78, another ER molecular chaperone, in the emotional brain regions such as the prefrontal cortex (PFC), hypothalamus, hippocampus and amygdala. Comprehensive behavioral analyses revealed that Atf6b-/- mice exhibited anxiety-like behavior in the light/dark transition test and hyperactivity in the forced swim test. Consistently, PFC and hypothalamic corticotropin-releasing hormone (CRH) expression was increased in Atf6b-/- mice, as was circulating corticosterone. Moreover, CRH receptor 1 antagonism alleviated anxiety-like behavior in Atf6b-/- mice. These findings suggest ATF6β deficiency elicits anxiety-like behavior and hyperactivity in CRH receptor 1-dependent mechanism. ATF6β could play a role in psychiatric conditions in the emotional centers of the brain.

Neuropathic pain is a debilitating chronic syndrome of the nervous system caused by nerve injury. In Drosophila, the Hedgehog (Hh) signaling pathway is related to increased pain sensitivity (hyperalgesia) but does not affect the baseline nociceptive threshold. In general, the contribution of the Hh signaling pathway to neuropathic pain in vertebrates is a highly debated issue. Alternatively, we investigated the potential role of Hh signaling in mechanical allodynia using a mouse model of neuropathic pain. Seven days after spinal nerve-transection (SNT) surgery, microglial activation increased in the ipsilateral spinal dorsal horn compared with that in the sham group; however, 21 days after surgery, microglial activation decreased. Contrastingly, astrocyte activation in the spinal cord did not differ between the groups. On day 21 of postsurgery, the SNT group showed marked upregulation of sonic hedgehog expression in peripheral glial cells but not in dorsal root ganglion (DRG) neurons. Intrathecal administration of the Hh signaling inhibitor vismodegib attenuated the mechanical allodynia observed on day 21 postsurgery. Conversely, intrathecal treatment with the Hh signaling activator smoothened agonist in naive mice induced mechanical allodynia, which was abolished by the ATP transporter inhibitor clodronate. Moreover, inhibition of Hh signaling by pretreatment with vismodegib significantly reduced ATP secretion and the frequency/number of spontaneous elevations of intracellular calcium ion levels in cultured DRG cells. Thus, the Hh signaling pathway appears to modulate the neural activity of DRG neurons via ATP release, and it plays an important role in sustaining mechanical allodynia and hypersensitivity in a mouse model of neuropathic pain.

BACKGROUND Neuropathic pain (NeP) medications have several side effects that affect NeP patients' quality of life. Betanin, the most common betacyanin pigment, has been shown to have potent antioxidant and anti-inflammatory properties in vivo; thus, it has potential as a healthcare treatment. In this study, we focused on betanin (red beetroot extract) as a potential therapy for NeP. METHODS Mice model of NeP were made by chronic constriction injury (CCI), and the development of mechanical hypersensitivity was confirmed using the von Frey test. Motor coordination and locomotor activity were assessed using open field tests and rotarod tests, respectively. The expression level of glial markers in the spinal cords was analyzed by immunostaining. The direct effects of betanin on microglial cells were investigated using primary cultured microglial cells. RESULTS In CCI model mice, repeated betanin treatment, both intraperitoneally and orally, attenuated developing mechanical hypersensitivity in a dose-dependent manner without impairing motor coordination. Betanin treatment also attenuated mechanical hypersensitivity that had developed and prevented the onset of mechanical hypersensitivity in CCI mice. Microglial activation in the spinal cord is known to play a key role in the development of NeP; betanin treatment reduced CCI-induced microglial activation in the spinal cord of model mice. Moreover, in primary microglia cultured cells, the activation of microglia by lipopolysaccharide application was suppressed by betanin treatment. CONCLUSION Betanin treatment appears to ameliorate mechanical hypersensitivity related to CCI-induced NeP in mice by inhibiting microglial activation. SIGNIFICANCE This article supports findings of the effect of betanin on NeP and provides a potential therapeutic candidate for NeP. Furthermore, elucidating the underlying mechanism of the effect of betanin on microglial activation could assist the development of new treatments for chronic pain.