Rabbit lacrimal acinar cells in primary culture: Morphology and acute responses to cholinergic stimulation

Purpose. The rabbit lacrimal gland yields large numbers of viable acinar cells that, when exposed to carbachol, respond with accelerated protein release, fluid phase endocytosis (Lucifer yellow uptake), and Na/H antiport activation. The current study was undertaken to determine whether such cells exhibit similar responses after having been maintained in primary culture. Methods. Cells were isolated from 2-kg, juvenile male New Zealand White rabbits and maintained in a supplemented DMEM/Ham's F-12 medium for up to 72 hours. Results. Electron microscopy showed the organization of freshly isolated cells to be highly polarized, with secretory vesicles at one pole and nucleus at the other; vesicles were heterogeneous in size and in the electron density of their contents. The cells remained polarized after overnight culture, but the secretory vesicle population was more homogeneous in size and content, and the cells tended to aggregate. After 72 hours, roughly half the cells retained good morphology and cytoplasmic polarization, but the vesicles were enlarged and their contents less electron dense. Cells that had been maintained overnight responded to the addition of 10 μM carbachol with a 32.2% ± 15.5% (n = 12, P < 0.04) increase in the total amount protein released during a standard 20-minute incubation. This represented a mean 125% increase in the temperature-dependent component of protein release. The protein secretory response was decreased to 14.6% ± 6.1% (n = 3, P < 0.07) for cells that had been maintained for 72 hours. In the same samples, carbachol increased fluid phase endocytosis by 38.3% ± 8.1% (P < 0.01) and 70.9% ± 13.4% (P < 0.025), respectively. The protein secretory response was partially, and the endocytic response fully blocked by 1 mM atropine. Conclusions. This model could be useful as a simplified system in which to study regulation of acinar cell function over days, rather than hours, as is required in fresh tissue models.