Multidisciplinary Approach
In LaNEF, a variety of techniques are used to study neurobiology and the structure of its elements, including gene modification, cell electrophysiology, neuropharmacology, time-resolved fluorescence, and mimetic systems of membranes. These techniques are used to investigate the functions and mode of action of the endocannabinoid ligands, intracellular pH, channelopathies and ion channel gating, and cationic peptide neural action.
Molecular Biology
Molecular biology provides a series of useful techniques for manipulating a nucleotide sequence and checking protein expression. Specific gene can be selected and isolated. Molecular biology is specially useful for making single amino acid changes in neural proteins such as ion channels, ion transporters, and receptors.
DNA electrophoresis apparatus.
Agarose gel for DNA.
Whole Cell Voltage Clamp Recordings
A glass pipette filled with a solution that mimiks the intracellular medium is used to break the cell membrane. This procede creats an electrical access to the cell interior. Using specific-controlled electrical stimulus, changes in membrane potential or membrane current can be fine monitored via the patch recording pipette. Pharmacological agents can also be delivered to the cell by adding directly to the bathing solution (i.e., outside the cell) or into the cell via the patch pipette (i.e., inside the cell).
The patch clamp rig.
The patch clamp rig.
Kir2.6 channel current activity and its modification by TPP single mutations.
Kir2.6 channel current activity and its modification by TPP single mutations.
Mimetic Systems of Membranes
The cell membranes are a complex system formed by lipids and proteins. Each constituent adds into the membranes specific physical-chemistry properties which can alter the dynamic function of many classes of membrane proteins such as the ion transporters and channels. Mimetic systems of membranes are used to simplify the complexity of the cell membrane and to study the specific function of the lipids and membrane proteins. In LaNEF, planar lipid bilayers are used to obtain electrical information of protein-free or protein-incorporated membranes. Large unilamellar vesicles (LUV) are also used to perform structural and fluorimetric analysis of peptide-incorporated membranes.
The planar lipid bilayer rig.
Single-channel current activity of Gramicidin channel and its modulation by endocanabinoid ligands.
