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Study of sperm ion channels by Patch clamp (CAT#: STEM-PET-0027-WXH)

Introduction

Upon ejaculation, mammalian spermatozoa have to undergo a sequence of physiological transformations within the female reproductive tract that will allow them to reach and fertilize the egg. These include initiation of motility, hyperactivation of motility and perhaps chemotaxis toward the egg, and culminate in the acrosome reaction that permits sperm to penetrate the protective vestments of the egg. These physiological responses are triggered through the activation of sperm ion channels that cause elevations of sperm intracellular pH and Ca2+ in response to certain cues within the female reproductive tract.




Principle

The patch-clamp technique involves a glass micropipette forming a tight gigaohm seal with the cell membrane. The micropipette contains a wire bathed in an electrolytic solution to conduct ions. To measure single ion channels, a “patch” of membrane is pulled away from the cell after forming a gigaohm seal.

Applications

• Study of ionic currents in individual isolated living cells, tissue sections, or patches of cell membrane.
• Study of excitable cells such as neurons, cardiomyocytes, muscle fibers, and pancreatic beta cells.
• Study of ion channels.

Procedure

1. Fabrication of glass electrodes
2. Measuring glass electrode resistance and compensating offset potential
3. Glass electrode contact to cell membrane and obtain a GΩ seal
4. Acquire and analyse recordings using the appropriate software.

Materials

Patch clamp system