Atomic force microscopy in studies of the cochlea

Area of Science:

• Biophysics
• Cell Biology
• Auditory Neuroscience

Background:

• Mammalian hearing sensitivity relies on cochlear amplification, driven by outer hair cell (OHC) motility.
• OHC motility is mediated by a membrane protein called prestin, crucial for sound amplification.
• Understanding prestin's structure and function is vital for auditory research.

Purpose of the Study:

• To present a novel atomic force microscopy (AFM) method for observing prestin in cellular membranes.
• To visualize prestin's potential structural contribution to OHC function.
• To characterize prestin distribution in Chinese hamster ovary (CHO) cells.

Main Methods:

• Utilized AFM to image prestin expressed in transfected CHO cells under liquid conditions.
• Employed ultrasonic waves to prepare inside-out plasma membrane samples attached to a substrate.
• Immunostaining with anti-FLAG antibodies and fluorescence labeling of membrane lipids were performed.

Main Results:

• AFM imaging revealed distinct particle-like structures (8-12 nm diameter) in prestin-transfected CHO cells.
• These structures were significantly more abundant in cells expressing prestin compared to untransfected controls.
• The observed structures are hypothesized to be composed of prestin molecules.

Conclusions:

• The developed AFM method allows for the visualization of prestin in cellular membranes.
• The presence of prestin correlates with specific particle-like structures, potentially representing functional units.
• This technique offers a new avenue for studying prestin's role in auditory function at the molecular level.