![]() Optogenetic techniques have yet to offer anything comparable and are unlikely to ever be able to. No other technique gives experiments such precise control over membrane potentials. Also critically patch-clamp offers the ability to apply a voltage clamp or a current clamp to the membrane in order to best characterize the neurons electrical properties. By contrast the best temporal resolution offered by optogenetic recording techniques is on the order of tens of milliseconds, and due to optical interference can only be used on structure close to the surface or require removal of intervening structures. While other modern recording techniques such as multi-electrode array and fluorescent calcium sensors have emerged as alternatives which can record from a great deal more neurons at once, patch clamp offers millisecond temporal resolution and the ability to study specific ionic currents. Patch clamp in comparison to other electrophysiological recording techniques Patch clamp can be used to record from adjacent and nearby cells to understand how small cliques of neurons connect to each other and study sub-threshold events. Eventually these techniques were extended into awake, head-fixed animals and even freely moving rodents. The chance of the pipette tip clamping off target was reduced via application of positive pressure during insertion or use of a cleaning pipette to ensure the main seal was not damaged. The first in vivo recording was accomplished in anesthetized head fixed cats. Eventually these technical challenges were overcome. As with brain slice preparations off target physical barriers present an issue as well, and one that cannot be removed with enzymatic solutions. ![]() Achieving this with the delicate glass tubes with incredibly thin points extending into a tip used for patch-clamp requires significant mechanical stabilization either by head fixing the animal or use of a stabilization rig attached to the animals head. Neuroscience often requires concordant observation of electrophysiological dynamics and behavior to test the relationship between psychological theories and their underlying biology. In time it was found that application of enzymatic solutions enabled recordings in brain slices as well. Brain slice preparations have debris, blood and lymph vessels or off target cells which may block the pipette from its target. Initially the need for direct contact between the pipette and the cell membrane limited patch clamp's use to cultured cell preparations. Many medical conditions are the result of a channelopathy, a mutation of a gene coding for an ion channel which disrupts electrical phenomenon underlying physiological processes. Characterizing the opening dynamics of ion channels has provided crucial insights to physiological mechanisms underlying a diverse set of conditions from the neurological, to diabetes, and heart conditions. Both Erwin Neher and Bert Sakmann, the inventors of the technique, went on to win the 1991 Nobel Prize in Physiology or Medicine for developing patch clamp and using it to prove the existence of ion channels. The pipette solution can also be filled with specific ions in order to target particular ion channels of interest such as sodium channels, potassium channels, and calcium channels among many others. This clamp is critical for testing voltage dependent opening dynamics of ion channels which determine a membrane's resting potentials and action potentials. This ability to form the voltage clamp on a patch of membrane give the technique its name. The high resistance seal allows the experimenter to hold the patch of membrane within the seal at a desired voltage for study of voltage-gated ion channels. ![]() While this seal will be of the order of megaohms, it has been found that applying slight suction will result in a seal with a resistance greater than a giga-ohm. The pipette is filled with a salt bath to conduct ionic currents before being pressed onto a cells surface to form an electrical seal with a high electrical resistance (measured in the unit ohm) which ensures low noise in the recording. Modern preparations use different tip diameters depending on the intended application. The technique was originally developed using a small glass tube, pipette, which is rapidly heated and stretched to produce a needle like shape with an opening diameter of 3-5 millimeters. ![]() Patch-seq is a specialized form of the patch clamp recording technique, the gold standard of single cell electrophysiological studies for its millisecond resolution of cellular electrophysiology, its ability to detect currents of specific ions, and perhaps most importantly for its ability to form a voltage clamp on the cell membrane. A depiction of a pipette forming a seal with a neuronal soma, negative pressure is applied to form the giga-ohm seal ![]()
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