Biophysics: Channelopathies

• This is a way of recording from neurons without destroying the cell
• A patch pippette forms a high resistance giga-ohm seal with the cell, creating an on cell patch.
• Negative pressure is then used to **** a bit of the membrane into the pippette, with the inside facing outward, creating an inside out patch. Good for studying 2nd messenger systems.
• Outside out is made from a whole cell clamp and is the reverse it is good for studying ion channels.

Cell-attached patch recordings

• A type of patch clamping.
• Quick inward current characteristic of an Na channel, slow outward current characteristic of a K+ channel.

• Na channels open, and then are inactivated using the inactivation gate, before properly closing.
• K+ channels only have open and closed states. 

• A gating current is the small current that moves across voltage gated channels upon voltage application
• It is so small that it can only be detected by removing the usual currents that the channel produces using TTX to stop Na channels, or caesium to stop K channels.

• The two main types are voltage gated and ligand gated.
• Voltage gated channels include Na, K, and Ca channels.
• Ligand gated include the glutamate receptors.

BK channels

• These are opened by both voltage and intracellular Ca, a combination fo which gives the largest opening. 

Sodium channels

• Four repeats of six transmembrane domains.
• The alpha subunit contains the pore  (has the four repeats).
• TTX binds to sodium channels, but Glutamate->glycine stops this interaction (between s5 and s6 of repeat 1). 
• A ring of negative charge is how the channel selects for posivie charge 
• S4 is the voltage sensor and has lots of positively charged residues
•              It is an alpha helix and membrane potential changes induce conformational changes in               it
•             When you reduce the positive charge of S4, the gating current is reduced.

Potassium channels

• 4 proteins of six transmembrane domains, which can be heteromultimeric.

• Follows the hinged lid model.
• It uses isoleucine, phenylalanine and methionine between repeats 3 and 4
•                    They form a lid to inactivate the channel
• If you remove the phenylalanine, the channel cannot be inactivated, and shows behaviour more like potassium channels.

Paramyotonia congenita

• It exhibits dominant inheritence and causes muscle stiffness preceded by paralysis
• S4 of repeat 4 is mutated, which usually couples activation with inactivation

Hyperkalemic periodic paralysis

• Once again, attacks of muscle weakness triggered by heightend blood potassium levels (4 to 5 mM).
• Incomplete inactivation of the Na channel

Potassium-aggravated myotonia

• Aggravated by potassium rich food
• Mutations in the inactivation loop, whereby a glycine is replaced by a glutamate
• Causes a slower closure of Na currents. 

• The presence or absence of paralysis depends on the fraction of Na current that doesn't turn off