Direct Current Stimulation Disrupts Endothelial Glycocalyx and Tight Junctions of the Blood-Brain Barrier in vitro.

Transcranial direct current stimulation (tDCS) is a non-invasive physical therapy to treat many psychiatric disorders and to enhance memory and cognition in healthy individuals. Our recent studies showed that tDCS with the proper dosage and duration can transiently enhance the permeability (P) of the blood-brain barrier (BBB) in rat brain to various sized solutes. Based on the in vivo permeability data, a transport model for the paracellular pathway of the BBB also predicted that tDCS can transiently disrupt the endothelial glycocalyx (EG) and the tight junction between endothelial cells. To confirm these predictions and to investigate the structural mechanisms by which tDCS modulates P of the BBB, we directly quantified the EG and tight junctions of in vitro BBB models after DCS treatment. Human cerebral microvascular endothelial cells (hCMECs) and mouse brain microvascular endothelial cells (bEnd3) were cultured on the Transwell filter with 3 µm pores to generate in vitro BBBs. After confluence, 0.1-1 mA/cm2 DCS was applied for 5 and 10 min. TEER and P to dextran-70k of the in vitro BBB were measured, HS (heparan sulfate) and hyaluronic acid (HA) of EG was immuno-stained and quantified, as well as the tight junction ZO-1. We found disrupted EG and ZO-1 when P to dextran-70k was increased and TEER was decreased by the DCS. To further investigate the cellular signaling mechanism of DCS on the BBB permeability, we pretreated the in vitro BBB with a nitric oxide synthase (NOS) inhibitor, L-NMMA. L-NMMA diminished the effect of DCS on the BBB permeability by protecting the EG and reinforcing tight junctions. These in vitro results conform to the in vivo observations and confirm the model prediction that DCS can disrupt the EG and tight junction of the BBB. Nevertheless, the in vivo effects of DCS are transient which backup its safety in the clinical application. In conclusion, our current study directly elucidates the structural and signaling mechanisms by which DCS modulates the BBB permeability.

Modulation of solute diffusivity in brain tissue as a novel mechanism of transcranial direct current stimulation (tDCS).

The breadth of brain disorders and functions reported responsive to transcranial direct current stimulation (tDCS) suggests a generalizable mechanism of action. Prior efforts characterized its cellular targets including neuron, glia and endothelial cells. We propose tDCS also modulates the substance transport in brain tissue. High resolution multiphoton microscopy imaged the spread across rat brain tissue of fluorescently-labeled solutes injected through the carotid artery after tDCS. The effective solute diffusion coefficient of brain tissue (Deff) was determined from the spatio-temporal solute concentration profiles using an unsteady diffusion transport model. 5-10 min post 20 min-1 mA tDCS, Deff increased by ~ 10% for a small solute, sodium fluorescein, and ~ 120% for larger solutes, BSA and Dex-70k. All increases in Deff returned to the control level 25-30 min post tDCS. A mathematical model for Deff in the extracelluar space (ECS) further predicts that this dose of tDCS increases Deff by transiently enhancing the brain ECS gap spacing by ~ 1.5-fold and accordingly reducing the extracellular matrix density. The cascades leading ECS modulation and its impact on excitability, synaptic function, plasticity, and brain clearance require further study. Modulation of solute diffusivity and ECS could explain diverse outcomes of tDCS and suggest novel therapeutic strategies.

In Vivo Modulation of the Blood-Brain Barrier Permeability by Transcranial Direct Current Stimulation (tDCS).

tDCS has been used to treat various brain disorders and its mechanism of action (MoA) was found to be neuronal polarization. Since the blood-brain barrier (BBB) tightly regulates the neuronal microenvironment, we hypothesized that another MoA of tDCS is direct vascular activation by modulating the BBB structures to increase its permeability (P). To test this hypothesis, we used high resolution multiphoton microscopy to determine P of the cerebral microvessels in rat brain. We found that 20 min 0.1-1 mA tDCS transiently increases P to a small solute, sodium fluorescein (MW 376) and to a large solute, Dextran-70k, with a much higher increase in P to the large solute. By pretreating the vessel with a nitric oxide synthase inhibitor, we revealed that the tDCS-induced increase in P is NO dependent. A transport model for the BBB was further employed to predict the structural changes by the tDCS. Comparing model predictions with the measured data suggests that tDCS increases P by temporarily disrupting the structural components forming the paracellular pathway of the BBB. That the transient and reversible increase in the BBB permeability also suggests new applications of tDCS such as a non-invasive approach for brain drug delivery through the BBB.