Phar0011: Molecular Pharmacology 2024-2025C/C++

Java Python Phar0011: Molecular Pharmacology

2024-2025

Level-6 Students: Sodium channel block summative data analysis exercise.

In this exercise the effect is studied of two blockers of sodium channels, tetrodotoxin and procaine, on the compound action potential recorded extracellularly from an isolated frog sciatic nerve. The aim of the exercise is to test the hypothesis that the two blockers act on the same channel site: this is done by measuring their effect when applied individually and then simultaneously (see Theory appendix).

Tetrodotoxin (TTX) is a powerful paralytic toxin which is found in Tetraodontid fish, including puffer fish, the Japanese fugu.  It has almost no therapeutic use, but is a valuable tool in experimental neurophysiology.

Procaine is a synthetic agent which was used clinically as a local anaesthetic.

The APPENDIX outlines the theory needed to predict the proportion of channels blocked by TTX or procaine or a mixture of the two.

Draw a diagram of a voltage-dependent sodium channel and label the diagram with the main functional domains of the channel.  Indicate on your diagram the regions ofTTX and procaine binding.

METHOD

Frog sciatic nerve was used to make the measurements to be analysed in this exercise. Briefly, the

sciatic nerve was dissected from apithed frog and mounted in a recording chamber, with stimulating electrodes at the distal (knee) end of the nerve and recording electrodes at the proximal end. Typical stimulation parameters are voltage pulses of 0.1 ms duration at a frequency of 0.1 s-1.   The stimulus strength was adjusted to twice the voltage required to give the maximal compound action potential amplitude. The central section of the nerve was cleaned of its connective tissue sheath in order to allow the blockers a faster access to axonal membranes.  This de-sheathed portion of the nerve was in the centre compartment of the nerve bath and application of drug solutions was restricted to this compartment by vaseline seals around the nerve that prevent drugs from reaching the stimulating and recording chambers.

EXPERIMENTAL RESULTS

i) Control action potential. The compound action potential recorded with an extracellular electrode has a biphasic waveform.

Question 1. Why is the compound action potential biphasic?

ii) The action potential conduction velocity was measured to be 28 ms-1.

Question 2. What does this tell you about the types of nerve fibres which contribute to the action potential in frog sciatic