Human red cell AQP1 is the first functionally defined member of the aquaporin family of membrane water channels. Here we describe an atomic model of AQP1 at 3.8A resolution from electron crystallographic data. Multiple highly conserved amino-acid residues stabilize the novel fold of AQP1. The aqueous pathway is lined with conserved hydrophobic residues that permit rapid water transport, whereas the water selectivity is due to a constriction of the pore diameter to about 3 A over a span of one residue. The atomic model provides a possible molecular explanation to a longstanding puzzle in physiology-how membranes can be freely permeable to water but impermeable to protons.

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.

Membrane water transport is critically involved in brain volume homeostasis and in the pathogenesis of brain edema. The cDNA encoding aquaporin-4 (AQP4) water channel protein was recently isolated from rat brain. We used immunocytochemistry and high-resolution immunogold electron microscopy to identify the cells and membrane domains that mediate water flux through AQP4. The AQP4 protein is abundant in glial cells bordering the subarachnoidal space, ventricles, and blood vessels. AQP4 is also abundant in osmosensory areas, including the supraoptic nucleus and subfornical organ. Immunogold analysis demonstrated that AQP4 is restricted to glial membranes and to subpopulations of ependymal cells. AQP4 is particularly strongly expressed in glial membranes that are in direct contact with capillaries and pia. The highly polarized AQP4 expression indicates that these cells are equipped with specific membrane domains that are specialized for water transport, thereby mediating the flow of water between glial cells and the cavities filled with CSF and the intravascular space. Key words: aquaporin-4 water channel; brain water permeability; glial cells; ependymal cells; immunogold electron microscopy; CSFWater metabolism is of major importance in a number of physiological processes in the CNS including CSF production and absorption, fluid transport across neuropil and vascular endothelium, and cell volume regulation (Fitzsimons, 1992;Robertson, 1992). In addition, water transport may serve to compensate for local changes in osmolality associated with potassium siphoning, which is essential for synaptic transmission. Alterations in water distribution in brain and CSF compartments is a common occurence in multiple neuropathological conditions including brain edema, brain tumors, stroke, hyponatremia, head injuries, and hydrocephalus. Despite its importance, little is known about the cellular and molecular mechanisms involved in transmembrane water movements in brain.Discovery of aquaporin-1 (Preston et al., 1992) answered the long-standing biophysical question of how water crosses plasma membranes (for review, see Agre et al., 1993;Knepper, 1994). Characterization of aquaporins provided molecular insight into fundamental processes of normal water balance and disorders of water balance outside brain (for review, see Nielsen et al., 1996). A cDNA for aquaporin-4 (AQP4) water channel protein was isolated recently from rat brain (Hasegawa et al., 1994;Jung et al., 1994), and abundant AQP4 was noted in brain including in cerebellum, hypothalamus, spinal cord, and ependymal cells lining the ventricles (Jung et al., 1994;Frigeri et al., 1995). Nevertheless, the cellular and subcellular distributions of AQP4 in brain remain unknown, and definition of the sites of AQP4 expression will be essential for understanding its physiological and pathophysiological roles.Immunocytochemistry and high-resolution immunogold electron microscopy were used to define the sites of AQP4 in brain. AQP4 expression is restricted to ependymal cell lining of ...