Stimulated Transepithelial Ion Flux in a Choroid Plexus Epithelial Cell Line

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Production and maintenance of the cerebral spinal fluid (CSF) is crucial for brain health and normal function. Alterations in normal homeostatic balance can occur in response a variety of pathological etiologies including traumatic brain injury, stroke, infection and cancerous lesions. Overproduction or blockage of CSF flow can also lead to hydrocephalus in both the pediatric population and adults. These alterations are remarkably difficult to treat and, in many cases, surgery is the only option. The choroid plexus, an epithelial cell lined tuft of capillaries that project into the cerebral ventricles, is responsible for a substantial portion of the CSF production. The capillaries of the CP are relatively leaky while the epithelial cells surrounding this vasculature form the blood‐CSF barrier and control the amount and composition of the CSF. A stable porcine choroid plexus epithelial cell line (PCP‐R) with a high transepithelial resistance (TEER) has been developed. We used Ussing style electrophysiology to characterize stimulated transepithelial ion transport measured as short circuit current (SCC) in confluent PCP‐R cells. In response to forskolin, a compound known to activate adenylyl cyclase and increase intracellular cAMP, there is a sustained increase in transepithelial ion flux that is consistent with an increase in cation absorption and/or anion secretion. This stimulated transport is partially inhibited by 1 mM BaCl2, used as a pan inhibitor of K+ channels. It is also partially inhibited by 10 mM T16Ainh‐A01, an inhibitor of the Ca2+ activated Cl− channel TMEM16a and by 50 nM GLY H101, an inhibitor of the cAMP‐stimulated Cl− channel CFTR. Neither the stimulatory nor inhibitory responses were accompanied by a substantial change in TEER. An agonist of the transient receptor potential vanilloid 4 (TRPV4) cation channel, GSK1016790, elicits a prolonged ion transport response that is the opposite of the forskolin stimulated response. The TRPV4 agonist elicits an increase in cation secretion and/or anion absorption. Interestingly, this response to the agonist is accompanied by a reversible decrease in TEER (>50%). The effect of the agonist on both SCC and TEER can be blocked by pretreatment with HC067047, a TRPV4 antagonist. If added after agonist stimulation, the TRPV4 antagonist can also fully reverse the agonist stimulated ion flux and can substantially restore the agonist stimulated decrease in TEER. Studies such as these are important for characterizing the factors that may disturb the normal CSF production and lead to an accumulation of CSF that can, over time, cause cellular loss and brain injury.Support or Funding InformationThis research was supported by an Indiana University Collaborative Research Grant fund of the Office of the Vice President for Research, and by a grant from the Vice Chancellor's Office, IUPUI.