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Lens fiber cell gap junctions contain [[alpha].sub.3] (Cx46) and [[alpha].sub.8] (Cx50) connexins. To examine the roles of the two different connexins in lens physiology, we have genetically engineered mice lacking either [[alpha].sub.3] or [[alpha].sub.8] connexin. Intracellular impedance studies of these lenses were used to measure junctional conductance and its sensitivity to intracellular pH. In Gong et al. (1998), we described results from [[alpha].sub.3] connexin knockout lenses. Here, we present original data from [[alpha].sub.8] connexin knockout lenses and a comparison with the previous results. The lens has two functionally distinct domains of fiber cell coupling. In wild-type mouse lenses, the outer shell of differentiating fibers (see Fig. 1, DF) has an average coupling conductance per area of cell-cell contact of ~1 S/[cm.sup.2], which falls to near zero when the cytoplasm is acidified. In the inner core of mature fibers (see Fig. 1, MF), the average coupling conductance is ~0.4 S/[cm.sub.2], and is insensitive to acidification of the cytoplasm. Both connexin isoforms appear to contribute about equally in the DF since the coupling conductance for either heterozygous knockout (+/-) was ~70% of normal and 30-40% of the normal for both -/- lenses. However, their contribution to the MF was different. About 50% of the normal coupling conductance was found in the MF of [[alpha].sub.3] +/- lenses. In contrast, the coupling of MF in the [[alpha].sub.8] +/- lenses was the same as normal. Moreover, no coupling was detected in the MF of [[alpha].sub.3] -/- lenses. Together, these results suggest that [[alpha].sub.3] connexin alone is responsible for coupling MF The pH- sensitive gating of DF junctions was about the same in wild-type and [[alpha].sub.3] connexin -/- lenses. However, in [[alpha].sub.8] -/- lenses, the pure [[alpha].sub.3] connexin junctions did not gate closed in the response to acidification. Since [[alpha].sub.3] connexin contributes about half the coupling conductance in DF of wild-type lenses, and that conductance goes to zero when the cytoplasmic pH drops, it appears [[alpha].sub.8] connexin regulates the gating of [[alpha].sub.3] connexin. Both connexins are clearly important to lens physiology as lenses null for either connexin lose transparency. Gap junctions in the MF survive for the lifetime of the organism without protein turnover. It appears that [[alpha].sub.3] connexin provides the long-term communication in MF Gap junctions in DF may be physiologically regulated since they are capable of gating when the cytoplasm is acidified. It appears [[alpha].sub.8] connexin is required for gating in DF. KEY WORDS: pH gating * coupling conductance * [[alpha].sub.8] connexin Cx50 * [[alpha].sub.3] connexin Cx46

Fiber cells of the lens are interconnected by an extensive network of gap junctions containing [[Alpha].sub.3] (Cx46) and [[Alpha].sub.8] (Cx50) connexins. A specific role for these connexins in lens homeostasis is not known. To determine the contribution of these connexins to lens function, we used impedance techniques to study cell-to-cell coupling in lenses from homozygous [[Alpha].sub.3] knockout (-/-), heterozygous (+/-), and wild-type (+/+) mice. Western blots and immunofluorescence data indicated that [[Alpha].sub.8] remained at similar levels in the three classes of lenses, whereas [[Alpha].sub.3] was approximately 50% of the normal level in the +/- lenses, and it was absent from the -/- lenses. Moreover, the data from +/+ lenses suggest that a cleavage of connexins occurs abruptly between the peripheral shell of differentiating fibers (DF) and the inner core of mature fibers (MF). The appearance of the cleaved connexins was correlated to a change in the coupling conductance. In -/- lenses the coupling conductance of MF was zero, and these fibers were depolarized by about 30 mV from normal ([approximately equal to]-65 mV). The DF remained coupled, but the conductance was reduced to 30-35% of normal. However, the gap junctions in the DF of [[Alpha].sub.3] -/- lenses remained sensitive to pH. We conclude that [[Alpha].sub.3] connexin is necessary for the coupling of central fibers to peripheral cells, and that this coupling is essential for fiber cell homeostasis because uncoupled MF depolarize and subsequently become opaque.

Mutation of the alpha3 connexin gene in mice did not affect the initial formation of lenses and lens fiber differentiation but caused proteolytic degradation of crystallins in mice with a double dose of the mutant gene. This led to the development of nuclear cataracts. These results reveal the importance of gap junctions in the maintenance of lens transparency. Gap junctions are the means of adjacent cell to communicate with one another and may function as a structural component in cellular organization.

The enzyme oleamide hydrolase is responsible for the inactivation of neuromodulatory fatty acid amides, bioactive signalling molecules. The protein sequence of the fatty-acid amide hydrolase (FAAH) has similarities with the protein sequence of the various amidase enzyme sequences taken from divergent organisms. These different amidases share a common identifying sequence. The FAAH cDNA transfected COS-7 cells show high activity levels of oleamide hydrolase.

Cell communication is essential to maintain the organization of cells. It coordinates cellular activities while at the same time, lets cells retain their individuality. Direct cell-cell contact is achieved via specialized cell surface membrane structures called gap junctions. Gap junctions are made up of a family of proteins called connexins which define the properties of gap junction channels such as permeability to molecules and voltage sensitivity.

Laser scanning confocal microscopic analysis of epidermal gap junctions reveals that gap junctions are present as junctional plaques that composed of alpha(1) and beta(2) connexins during the embryonic development. Beta(2) connexin is selectively expressed in the upper and granular differentiated spinous layers. Alpha(1) is detected in the developing basal layer and less differentiated spinous layer. The segregated domains of both connexins encode a single homooligomer protein.