Molecular characterization of an aquaporin cDNA from brain: candidate osmoreceptor and regulator of water balance.

Abstract

The aquaporins transport water through membranes of numerous tissues, but the molecular mechanisms for sensing changes in extracellular osmolality and regulating water balance in brain are unknown. We have isolated a brain aquaporin by homology cloning. Like aquaporin 1 (AQP1, also known as CHIP, channel-forming integral membrane protein of 28 kDa), the deduced polypeptide has six putative transmembrane domains but lacks cysteines at the known mercury-sensitive sites. Two initiation sites were identified encoding polypeptides of 301 and 323 amino acids; expression of each in Xenopus oocytes conferred a 20-fold increase in osmotic water permeability not blocked by 1 mM HgCI2, even after substitution of cysteine at the predicted mercury-sensitive site. Northern analysis and RNase protection demonstrated the mRNA to be abundant in mature rat brain but only weakly detectable in eye, kidney, intestine, and lung. In situ hybridization of brain localized the mRNA to ependymal cells lining the aqueduct, glial cells forming the edge of the cerebral cortex and brainstem, vasopressin-secretory neurons in supraoptic and paraventricular nuclei of hypothalamus, and Purkinje cells of cerebellum. Its distinctive expression pattern implicates this fourth mammalian member of the aquaporin water channel family (designated gene symbol, AQP4) as the osmoreceptor which regulates body water balance and mediates water flow within the central nervous system. The aquaporins are a family of water-selective membrane channels found in animals, plants, and microorganisms (reviewed in refs. 1 and 2). Aquaporin 1 (AQP1, also known as CHIP, channel-forming integral membrane protein of 28 kDa) was the first protein shown to function as a molecular water channel (3) and is naturally expressed in mammalian red cells, renal proximal tubules (4-6), and other waterpermeable epithelia (7). AQP2 is the vasopressin-regulated water channel in renal collecting ducts (8, 9) and is the site of mutations in some forms of nephrogenic diabetes insipidus (10). AQP3 is the water channel in basolateral membranes of renal medullary collecting duct (11).Because of restricted space within the cranium, regulation of salt and water balance is essential for normal functions of the mammalian brain (reviewed in ref. 12). Moreover, vasopressin is released by the neurohypophysis in response to small changes in osmolality around the supraoptic and paraventricular nuclei (13). AQP1 (CHIP) is abundant in the choroid plexus, but not elsewhere in brain where molecular mechanisms responsible for transmembrane water movements and osmoreception are still unknown (7). By homology cloning we have isolated a cDNA from a rat brain library and have established its function and distribution. As this work was nearing completion, a highly related cDNA was cloned from rat lung (14); however, discrepancies with our studies were observed in the initiation site, in the coding and 3' untranslated sequences, and in the sites of expression and relative abundance. Thus, additional studie...

Identification of a novel protein containing two C2 domains selectively expressed in the rat brain and kidney

“…Whether B/K mediates similar functions in the kidney and the central nervous system remains to be determined. However, it is interesting that Jung et al recently isolated an aquaporin isoform from the brain and showed that it is expressed in vasopressin-secretory neurons in supraoptic and paraventricular nuclei of hypothalamus [25].…”

Cytoprotective effects of growth factors: BDNF more potent than GDNF in an organotypic culture model of Parkinson's disease

“…Cloning and molecular characterization of brain Aqp4 at the mRNA level was reported by two independent studies (18,297). The gene was reported to have two different initiation sites that gives rise to two functional proteins, one 301 and the other 323 amino acids long, corresponding to the two isoforms M23 (AQP4c) and M1 (AQP4a), respectively (18,297).…”

“…To date, eight aquaporins have been described in rodent brain cells, AQP1, AQP2, AQP3, AQP4, AQP5, AQP8, AQP9 and AQP11 (18,(259)(260)(261)(262)(263)(264)(265). Most of them are studied in cell cultures and only three of them are demonstrated in brain at both protein and mRNA level, namely AQP1, AQP4 and AQP9 (15,163).…”

“…The gene was reported to have two different initiation sites that gives rise to two functional proteins, one 301 and the other 323 amino acids long, corresponding to the two isoforms M23 (AQP4c) and M1 (AQP4a), respectively (18,297). When expressed in Xenopus oocytes, both the M1 and the M23 isoforms produce an increase in osmotic water flux identical to that of AQP1 (18,19), and when compared to other aquaporins, AQP4 shows the highest water permeability coefficient, followed by AQP1 (298). More recently, four new isoforms of AQP4 (AQP4b, d, e, f) have been described (299), in which one isoform (AQP4e) is expressed in astrocytic plasma membranes and mediate water transportation (300).…”

“…AQP4, on the other hand, is an orthodox aquaporin exclusively permeable to water. It is localized in astrocytic cell membranes and plays a crucial role in water-and osmoregulation, and hence in maintaining a homeostatic microenvironment required for neuronal function (18)(19)(20)(21). To date, however, the microenvironment surrounding the dopaminergic neurons in the SNpc has been largely neglected, although excess extracellular water in the SNpc has been demonstrated in patients with PD (22), thereby suggesting a role of AQP4 in PD.…”