טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
M.Sc Thesis
M.Sc StudentInataev Naila
SubjectMeasuring Ultrasound-Induced Nanoscopic Membrane
Deformations
DepartmentDepartment of Nanoscience and Nanotechnology
Supervisor Professor Emeritus Eitan Kimmel
Full Thesis textFull thesis text - English Version


Abstract

   During the last decades a rapid progress has been made in the scientific and clinical applications of invasive and non-invasive methods for stimulation of the nervous system. However, such techniques as electrode implants usually require surgical insertion and tissue displacement, typically becoming complex in biostability and biocompatibility issue. Recent studies also indicate that ultrasound technology, which is used extensively for medical imaging, is a suitable candidate for use in the field of neuro-modulation, yet the mechanism is not fully understood.

   A novel, physical model denoted as the "bilayer sonophore" (BLS), suggests that ultrasound waves cause a structural change in the membrane. The BLS model predicts that cellular membranes absorb ultrasound energy and inflate and deflate periodically whereas the distance between the two leaflets of the membrane increase and decrease respectively. In view of the BLS model, ultrasound activation of voltage-gated channels might be attributed to changes in the distance between the leaflets and subsequent changes in the membrane potential. This mechanism might explain the accumulating evidence that ultrasound can induce action potential in neuron cells.

   Here, we exposed various cell types including neurons and endothelial cells and cell-like liposomes (a micrometric spherical drop of liquid enclosed by an artificial bilayer membrane) to pulses of low intensity ultrasound in an attempt to study neural stimulation; and at the same time to investigate the periodic membranes swelling that was predicted by the BLS model predictions. Calcium imaging was used for measuring the electrical activity of primary cultures of rat cortical neurons, and fluorescence energy transfer (FRET) was used in neurons, endothelial cells and liposomes to demonstrate membrane deformation. The FRET method is sensitive to nanometric changes in the distance between two fluorescence molecules. We found that low intensity ultrasound induces a significant rise in the rate of generation of action potentials in neurons compared to control. Moreover, low intensity ultrasound was also found to stimulate action potentials in neurons treated first by ion channel blockers; and that the rate of generation of action potentials intensified even further when encapsulated gas microbubbles were introduced into the medium that surround the cells. As to direct validation of the BLS model, we observed for the first time ever, in a preliminary set of experiments, a significant BLS swelling in liposomes. Similar effects were observed in neurons in the low range of low intensity ultrasound but not in endothelial cells.