Water rapidly crosses the plasma membrane of red blood cells (RBCs) and renal tubules through specialized channels. Although selective for water, the molecular structure of these channels is unknown. The CHIP28 protein is an abundant integral membrane protein in mammalian RBCs and renal proximal tubules and belongs to a family of membrane proteins with unknown functions. Oocytes from Xenopus laevis microinjected with in vitro-transcribed CHIP28 RNA exhibited increased osmotic water permeability; this was reversibly inhibited by mercuric chloride, a known inhibitor of water channels. Therefore it is likely that CHIP28 is a functional unit of membrane water channels.

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...

Despite longstanding interest by nephrologists and physiologists, the molecular identities of membrane water channels remained elusive until recognition of CHIP, a 28-kDa channel-forming integral membrane protein from human red blood cells originally referred to as "CHIP28." CHIP functions as an osmotically driven, water-selective pore; 1) expression of CHIP conferred Xenopus oocytes with markedly increased osmotic water permeability but did not allow transmembrane passage of ions or other small molecules; 2) reconstitution of highly purified CHIP into proteoliposomes permitted determination of the unit water permeability, i.e., 3.9 x 10(9) water molecules.channel subunit-1 x s-1. Although CHIP exists as a homotetramer in the native red blood cell membrane, site-directed mutagenesis studies suggested that each subunit contains an individually functional pore that may be reversibly occluded by mercurial inhibitors reacting with cysteine-189. CHIP is a major component of both apical and basolateral membranes of water-permeable segments of the nephron, where it facilitates transcellular water flow during reabsorption of glomerular filtrate. CHIP is also abundant in certain other absorptive or secretory epithelia, including choroid plexus, ciliary body of the eye, hepatobiliary ductules, gall bladder, and capillary endothelia. Distinct patterns of CHIP expression occur at these sites during fetal development and maturity. Similar proteins from other mammalian tissues and plants were later shown to transport water, and the group is now referred to as the "aquaporins." Recognition of CHIP has provided molecular insight into the biological phenomenon of osmotic water movement, and it is hoped that pharmacological modulation of CHIP function may provide novel treatments of renal failure and other clinical problems.