Introduction: Cerebrospinal fluid (CSF) is produced predominately by the choroid plexus epithelium (CPe). The composition of CSF varies according to physiological, diurnal, and pathophysiological influences. Understanding the transporters that control the production and composition of CSF is of clinical relevance for many diseases. In a genetic rat model of hydrocephalus (Tmem67 ^-/-), we have previously shown that antagonists of the transient receptor potential vanilloid 4 (TRPV4) channel, ameliorate the development of excess CSF in the context of hydrocephalus, implicating this channel as a key component of CSF production. The aim of our studies is to use a human CPe cell model, the HIBCPP (human choroid plexus papilloma) to identify the transepithelial electrolyte fluxes that occur in response to TRPV4 stimulation and to determine if the electrolyte flux is accompanied by a measurable fluid movement.

Methods: HIBCPP cultures were grown on Millicell permeable supports. Ussing-style electrophysiology was used to define changes in transepithelial electrogenic fluxes (short circuit current, I_SC) and transepithelial permeability (conductance, the inverse of transepithelial resistance) in the presence of TRPV4 agonists and effectors.  Parallel cultures were used to measure fluid secretion (net fluid flux from the basolateral media to the apical media) or absorption (the opposite of secretion) 10 minutes after the addition of a TRPV4 agonist.

Results: We previously showed that the HIBCPP cell line has important transporters found in the native epithelium in the correct polarization and forms a moderately tight barrier epithelium consistent with the blood-CSF barrier. TRPV4 agonist stimulation causes a multicomponent change in transepithelial electrolyte flux and a substantial and reversible change in transepithelial permeability as measured by transepithelial conductance. The TRPV4-mediated electrolyte flux appears to be secondary activation of multiple transport proteins stimulated in response to the TRPV4-mediated influx of Ca²⁺ and Na⁺.  The I_SC is a complex mixture of movement of both cations and anions causing the I_SC to return to baseline within 10 minutes.  However, the increased conductance, which remains elevated at 10 minutes, indicates the I_SC return to baseline is a function of continuing but opposing transepithelial movements of electrolytes.  This flux is accompanied by a statistically significant fluid secretion. The vehicle-treated samples had an increase in fluid secretion of 1.34 μL/cm²/10 min (mean + SEM, n=5), a value not statistically different from zero. In response to the TRPV4 agonist, there was a significant (p=0.0297) 18.7-fold increase in fluid secretion to 25.11 μL/cm²/10 min (n=7). Although in vivo the choroid plexus is one of the most secretory epithelia in the body this is, to our knowledge, the only cultured epithelium showing this level of fluid secretion.

Conclusions:  The HIBCPP cell line has multiple characteristics of the native choroid plexus epithelium.  This line is being used to dissect pathways involved in CSF production and these will be presented.  Our results have uncovered unexpected levels of fluid movement in response to TRPV4 stimulation which can inform the role of opposing electrolyte movements and electroneutral transporters.