Arginine vasopressin-Venus reporter mice as a tool for studying magnocellular arginine vasopressin neurons.

Arginine vasopressin (AVP) synthesized in the magnocellular neurons of the hypothalamus is transported through their axons and released from the posterior pituitary into the systemic circulation to act as an antidiuretic hormone. AVP synthesis and release are precisely regulated by changes in plasma osmolality. Magnocellular AVP neurons receive innervation from osmosensory and sodium-sensing neurons, but previous studies showed that AVP neurons per se are osmosensitive as well. In the current study, we made AVP-Venus reporter mice and showed that Venus was expressed exclusively in AVP neurons and was upregulated under water deprivation. In hypothalamic organotypic cultures from the AVP-Venus mice, Venus-labeled AVP neurons in the supraoptic and paraventricular nuclei survived for 1 month, and Venus expression was upregulated by forskolin. Furthermore, in dissociated Venus-labeled magnocellular neurons, treatment with NaCl, but not with mannitol, decreased Venus fluorescence in the soma of the AVP neurons. Thus, Venus expression in AVP-Venus transgenic mice, as well as in primary cultures, faithfully showed the properties of intrinsic AVP expression. These findings indicate that AVP-Venus mice as well as the primary hypothalamic cultures could be useful for studying magnocellular AVP neurons.

Chemical chaperone 4-phenylbutylate reduces mutant protein accumulation in the endoplasmic reticulum of arginine vasopressin neurons in a mouse model for familial neurohypophysial diabetes insipidus.

Familial neurohypophysial diabetes insipidus (FNDI), characterized by progressive polyuria and loss of arginine vasopressin (AVP) neurons, is an autosomal dominant disorder caused by AVP gene mutations. Our previous studies with FNDI model mice demonstrated that mutant proteins accumulated in the endoplasmic reticulum (ER) of AVP neurons. Here, we examined therapeutic effects of the chemical chaperone 4-phenylbutylate (4-PBA) in FNDI mice. Treatment with 4-PBA reduced mutant protein accumulation in the ER of FNDI mice and increased AVP release, leading to reduced urine volumes. Furthermore, AVP neuron loss under salt loading was attenuated by 4-PBA treatment. These data suggest that 4-PBA ameliorated mutant protein accumulation in the ER of AVP neurons and thereby prevented FNDI phenotype progression.

Unfolded protein response in hypothalamic cultures of wild-type and ATF6α-knockout mice.

Recent studies suggest that endoplasmic reticulum (ER) stress in the hypothalamus could affect systemic homeostatic regulation in areas such as energy and water balance. Activating transcription factor 6α (ATF6α) is an ER stress transducer which increases the expression of ER chaperones and ER-associated degradation (ERAD) components under ER stress. In the present study, we examined the regulation of the unfolding protein response (UPR) in mouse hypothalamic cultures of wild-type (WT) and ATF6α(-/-) mice. Thapsigargin (TG), an ER stressor, significantly increased the mRNA expression of immunoglobulin heavy chain binding protein (BiP), spliced X-box binding protein 1 (XBP1), activating transcription factor 4 (ATF4), C/EBP homologous protein (CHOP), and ERAD components, in hypothalamic cultures of WT mice with the same threshold (0.1µM) and similar time courses. On the other hand, TG-induced upregulation of BiP and CHOP as well as most ERAD-related genes, but not spliced XBP1 or ATF4, was attenuated in ATF6α(-/-) mice compared with WT mice. Our data suggest that all the UPR arms are activated similarly in the mouse hypothalamus under ER stress conditions, where ATF6α regulates the expression of ER chaperones, CHOP, and ERAD components.