Action potential alterations induced by single neuron mechanical loading

Thesis

Since the 1940s, an extensive body of empirical evidence has been gathered about non-electrical alterations accompanying the voltage depolarisation of a neuron during the generation and propagation of an action potential (AP). The prevailing traditional electrochemical models do not account for these changes, and are as a result incomplete. At the same time, multiple research programmes have demonstrated how ultrasound (US) -a mechanical wave- can non-invasively and reversibly mechanically perturb neuronal functions. Strikingly, the mechanisms of action remain largely unknown.

In this framework, this thesis takes the approach to focus on single cell studies to better understand and exploit the multiphysics of the neuronal action potential and aims at studying the effects of direct mechanical loadings on the shape and dynamics of the neuronal signal. First, a multiphysics platform combining nanoindentation and patch clamp systems, assembled in an inverted microscope, is designed to simultaneously measure mechanical and electrophysiological properties of single neurons while imaging the cell morphology. The functionality of the setup is first used to provide a full characterisation of the neuronal cell line chosen here, i.e., dorsal root ganglion-derived cells, and then to quantify the alterations in the magnitude and dynamics of single neuron spontaneous APs before, during and after quasi-static and dynamic mechanical loadings over a wide range of frequencies (up to 1 MHz).

Quasi-static indentation tests show transiently smaller, wider and slower depolarising APs during compressions within the cell's linear viscoelastic regime, which translates in slower AP dynamics, as well as sustained hyperpolarised cells with longer repolarisation rates after the indentation. Dynamic loading results, instead, show narrower APs with shorter depolarisation and repolarisation phases, i.e., faster induced AP dynamics, at increasing frequencies, particularly for stimulations at around 1 MHz. In addition, results at this frequency are more evident after indentation, hinting at a cumulative or lagged effect of mechanical stimulation at US frequencies. These observations are attributed to the stiffening of the membrane and membrane cortex as a result of the mechanical oscillations, as well as potentially related phase state alterations. Taken together, these findings highlight the importance of mechanical cues in shaping the neuronal AP, and the often overlooked role of the membrane and its mechanical properties in determining cell functionality. In particular, the promoted faster AP dynamics that emerge after high frequency mechanical oscillations suggest a potential mechanism of US neuromodulation and support the assumption that direct mechanical cellular agitations induced by US stimulation protocols assist the observed neuromodulatory effects.

Authors: Tamayo Elizalde, M

• DOI: 10.5287/ora-z5ykpobav
• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford
• Language: English
• Keywords: neuronal action potential; ultrasound neuromodulation; patch-clamp; nanoindentation; neuron multiphysics; mechanoelectrical coupling; neuronal membrane mechanics
• Subjects: Action potentials (Electrophysiology); Cells--Mechanical properties