Modulation of Cerebrospinal Fluid Production by Transient Receptor Potential Vanilloid 4: Implications for Disorders of Brain Fluid Volume

Abstract

Cerebrospinal fluid (CSF) secretion into the brain ventricular system is an active process via the choroid plexus (CP) - a specialized convoluted epithelial sheet surrounding a fenestrated capillary bed which is fed from the cerebral arterioles. The final CSF composition differs from a simple plasma filtrate implicating the CP as an important regulatory site controlling CSF composition and volume. Flux across this blood-CSF barrier is tightly controlled by ion and water transport proteins. Transient Receptor Potential Vanilloid 4 (TRPV4) is a polymodal, non-selective cation channel which functions as a regulatory hub protein in epithelia, including the CP. Our previous studies have shown that TRPV4-selective antagonists were effective at reducing ventriculomegaly in a genetic rat model of severe postnatal hydrocephalus. In CP, we found that activation of TRPV4 caused the influx of both Ca2+ and Na+ ions, and entry of both of these ions was selectively blocked by a TRPV4 antagonist. Systemic treatment with TRPV4 antagonists reduced TRPV4 mRNA expression in the CP in hydrocephalic rats, but not in normal rats, indicating a role for TRPV4 as part of a regulatory pathway in CSF production. To study the mechanism of CSF secretion by the CP, we utilized two continuous, high resistance CP cell lines: the porcine choroid plexus – Reims (PCP-R) and the human choroid plexus papilloma (HIBCPP) cells. The current experiments utilize Ussing chamber electrophysiological techniques, western blotting, qRT-PCR, and fluorescent immunolabeling. Immunolabeling and mRNA expression demonstrate a robust abundance for TRPV4 and other well-characterized CP membrane proteins in both cell lines. In both lines, TRPV4 activation stimulates a large change in short circuit current (ISC), a measure of net electrogenic ion flux. This change in ISCis coupled with a substantial change in transepithelial membrane conductance, a measure of cellular permeability. The transepithelial membrane transport changes due to TRPV4 activation in human and porcine cells are similar in overall effect, but differ in baseline, duration, and magnitude, indicating species-specific CP transport mechanisms. In both cases, these responses can be inhibited by either pre- or post-incubation with TRPV4 antagonists, substantiating the specificity to TRPV4 activation. The human cells, but not the porcine cells, demonstrate a substantial baseline transepithelial transport which is dependent on K+, Na+, and Cl- gradients. Furthermore, in both the porcine and human cells, the TRPV4 response is dependent upon K+ secretion, Cl- absorption, and Na+ secretion. These data confirm a role for TRPV4 in regulating ionic gradients, modulating both electrogenic and electroneutral transporters, and controlling CSF production. The promising results of these studies, coupled with published data from other investigators suggesting a role for TRPV4 in blood-brain-barrier permeability and astrocyte volume regulation, shows the importance of TRPV4 as a regulatory hub protein in brain fluid volume regulation.