Typical reversal electrical profile traces are shown in
Figure 1 , where the electrical noise on the traces, using a PAG fragment for the reference microelectrode, was less than 0.2 mV. Electrical profile traces from each mouse investigated showed the same features as illustrated in
Figure 1 . The initial negative potential was the intracellular transmembrane potential of the squamous corneal epithelium. As the microelectrode was raised in 1-μm steps, a positive potential was recorded that remained at a similar value until the microelectrode went into open circuit. When the microelectrode returned on its downward traverse, the profile repeated well and showed a positive electrical potential of similar magnitude until the microelectrode reentered the epithelial cell. At this spatial resolution of 1 μm, the tear film generated a uniform positively charged profile with no detectable discontinuities. Profiles from 10 separate mouse tear films are shown in
Figure 2A . The average tear film thickness was 7.2 ± 0.7 μm (mean ± SD,
n = 10). In general, 9 of the 10 tear films were of similar thickness, and the remaining one, which was measurably thinner at 5 μm, also exhibited the lowest magnitude of the electrical potential. All the electrical profiles within the tear films showed a continuous and positive potential. Their mean potentials ± SEM were 4.7 ± 0.3, 9.6 ± 0.4, 5.6 ± 0.2, 13.5 ± 0.8, 4.6 ± 0.7, 3.6 ± 0.3, 2.7 ± 0.3, 2.8 ± 0.4, and 3.9 ± 0.3 mV. Individual electrical profiles appeared to increase their magnitude at the apical surface. Others showed the reverse tendency
(Fig. 2A) . On average
(Fig. 2B) , there was no significant gradient in the electrical profiles. There were, however, statistically significant differences in the average standing electrical potentials between different murine tear films. Similar profiles were obtained when the reference electrode was placed in a 1-μL drop of saline situated on the nasal quadrant of the mouse sclera. With these preparations, the recorded electrical profiles were not only much noisier, as would be expected on general principle,
17 but they also showed a much briefer period of stability of only a few minutes (described later). It was, on occasion, possible to collect reproducible data on exit and entry through the tear film thickness, but usually before the whole profile could be collected, the recording was lost. In these preparations, the initial average tear film thickness was measured at 7.5 ± 1.0 μm (mean ± SD,
n = 10) and typical standing electrical potentials varied between 7 and 14 mV. At the spatial resolution of 1 μm, we detected no region where the electrical potential was zero.