Effects of Electromagnetic Pulses on the Permeability of Blood-Brain-Barrier Gui-Rong Ding, Ph.D Department of Radiaition Medicine, Fourth Military Medical University, China

Introduction  EMP( Electromagnetic pulses) exposure resulted in various changes in morphology and function of nervous system.  BBB (Blood-Brain-Barrier) plays an important role in maintaining homeostasis in brain, and BBB disruption can initiate the development of brain injuries.  Our previous study indicate that membrane is one of the sensitive targets of EMP.

Blood–Brain Barrier and the Tight Junction Astrocytic Foot Process Tight junction Endothelium Basal Lamina

Our concern EMP exposure BBB openning ? Molecular mechanism?

Material and Methods SD rats, SD rats, male, 6-8 weeks old ， 200±20g Animals

EMP Generator GTEM-Cell Material and Methods Exposure apparatus

EMP parameters  Frequency band GHz  Half width100ns  Pulses number  Pulses number50, 100 and 200  Intensity , 200 and 400 kV/m  Intensity 50, 100, 200 and 400 kV/m Material and Methods

BBB permeability detection Serial frozen section, whole coronal slices of brain Evans Blue (EB) tracer and fluorescent microscope Lanthanum nitrate tracer and transmission electron microscope Protein expression Western blotting Immunofluorescence Material and Methods

200kV/m, 200pulses Results EMP induced BBB opening (dose-effect) 100kV/m, 200pulses 50kV/m, 200 pulses control

Results EMP induced BBB opening (dose-effect) 200kV/m, 50 pulses 200kV/m, 100pulses 200kV/m, 200pulses

Results 4h after exposure2h after exposure6 h after exposure 200kV/m, 200pulses EMP induced BBB opening (time-effect)

Results control (×10000) 200kV/m 200pulses (×10000) (×10000) EMP induced BBB opening

Changes of BBB tight junction related proteins after EMP exposure Control Occludin Claudin-5 ZO-1 3h after exposure (200kV/m, 200pulses)

Protein expression of tight junction related proteins after EMP exposure Con SOL CSK SOL CSK SOL CSK SOL CSK 0hr3hr6hr ZO-1 Occludin65kDa 220kDa Claudin-5 Con 0hr3hr 6hr Occludin65kDa 22kDa 220kDaZO-1 Membrane protein CSK SOL Cellular fractionation

Conclusions  EMP exposure may lead to an increase in the permeability of BBB in rats.  The injury of BBB tight junction was involved in EMP-induced BBB permeability increase.

Ongoing work Primary cultured brain microvascular endothelial cells(BMEC) Primary cultured brain astrocyte

In vitro BBB model TranswellTranswell Polyester- multiple- membrane TranswellTranswell AstrocytesAstrocytes Culture dish BMECBMEC AstrocytesAstrocytes TranswellTranswell

Blood Testicle Barrier

1h 2h control 6h Effects of EMP (200kV/m, 200 pulses) on the Permeability of Blood-Testicle -Barrier

Changes of BTB tight junction related protein-ZO-1 after EMP (200kV/m, 200 pulses) exposure control 1h 2h 6h

Further research  To set up in vitro BBB model  To identify the size of the molecules that get though BBB after different parameters EMP exposure  To clarify the molecular mechanism of EMP-induced BBB opening

Tight Junctions （ TJs ） BTB BRB BBB In common Barrier system EMP exposure Sensitive organs  By observing the permeability and changes of TJs in other “Barrier systems”, to find out the common mechanism of EMP-induced barrier injury. Further research