Perforated Patch Clamp: from pores to currents and the challenges in between

Patch clamping is a common electrophysiological technique used to study the electrical behaviour of single neurons in the brain. The most commonly used variety of patch-clamp is the whole-cell method, which involves forming a tight high-resistance seal onto the membrane of a cell and subsequently breaking the membrane with force to achieve a continuum between the patch pipette and the cell’s cytosolic solution, allowing for full electrical access. Whole-cell mode is useful for recording voltage or current responses from cells with a very low access resistance.

However, whole-cell patch clamp inevitably involves artificially manipulating the cell’s cytosolic composition as it mixes with the solution in the pipette. Pipettes are therefore normally filled with an “intracellular” solution that mimics the ionic composition inside the neurons they are targeting. However, the intracellular solution is a simplified version of the cell’s cytosolic contents, and mainly mimics the concentrations of the major ions that determine the resting membrane potential (sodium, potassium, and chloride). This often results in washout of key endogenous cytosolic signalling molecules, such as cAMP and ATPase, which could disrupt receptor-mediated downstream signalling pathways. Normal intracellular calcium buffering is also affected, as exogenous calcium buffers such as EGTA are normally added to the pipette solution. These cytosolic components that are prone to washout are especially crucial in synaptic plasticity events, which are among the most commonly studied using patch clamp. Thus, there is a need for a method that enables electrophysiological recordings without altering the cell’s cytosolic composition, but still achieves a relatively low access resistance.