Jose L. Mercado

 

 

E-mail: jlmercado@wisc.edu

Research Project

Gamma-Aminobutyric acid (GABA) mediates most of the inhibitory transmission events in the mammalian CNS via activation of ionotropic GABAA receptors (GABAARs). GABAARs are of particular interest, from a pharmacological point of view, since they are the target for a wide range of therapeutic agents such as benzodiazepines (BZDs), barbiturates and anesthetics. The proposed mechanistic framework for GABAAR activation suggests that binding of GABA to the binding site triggers a series of conformational changes in the receptor that leads to the opening of a chloride (Cl-) ion channel and desensitization. This process is known as receptor gating. Even though these processes have been extensively studied, the structural rearrangements that govern receptor gating are not well understood.

The overall goal of this thesis proposal is to investigate the structural determinants important for coupling GABA binding to channel activation. I propose to investigate this by using several approaches including the substituted cysteine accessibility method (SCAM), two electrode voltage clamping as well as patch clamp recording.

Abstracts and Publications:

• Venkatachalan, S.P., J.D. Bushman, J.L. Mercado, F. Sancar, K.R. Christopherson, and A.J. Boileau. 2007. Optimized expression vector for ion channel studies in Xenopus oocytes and mammalian cells using alfalfa mosaic virus. Pflügers Arch – 454: 155-163.
• Mercado, J.L. and C. Czajkowski. 2007. GABA and pentobarbtial induce different conformational rearrangements in the alpha1 and beta2 pre-M1 regions of the GABAA receptor. Biophysical Soc. Abstr.
• Mercado, J. and C. Czajkowski. 2006. Charged residues in the alpha1 and beta2 pre-M1 regions involved in GABAA receptor activation. J. Neurosci. 26: 2031-2040.
• Mercado, J.L. and C. Czajkowski. 2003. Charged residues in the Pre-M1 region of the GABAA receptor play a role in coupling binding to gating. Soc. Neurosci. Abstr.
• Cruz-Martín, A., J.L. Mercado, L.V. Rojas, M.G. McNamee, and J.A. Lasalde-Dominicci. 2001. Tryptophan substitutions at lipid-exposed positions of the gamma M3 transmembrane domain increase the macroscopic ionic current response of the torpedo californica nicotinic acetylcholine receptor. J. Membrane Biol. 183: 61-70. [PDF]
• Mercado, J.L., A. Cruz-Martín, L.V. Rojas, M. McNamee, and J.A. Lasalde-Dominicci. 2001. Tryptophan substitutions in the gamma M3 transmembrane domain alter allosteric transitions of the torpedo californica nicotinic acetylcholine receptor. RISE-MBRS Abstr.
• Mercado, J.L., A. Cruz-Martín, L.V. Rojas, M. McNamee, and J.A. Lasalde-Dominicci. 2001. Hydrophobic amino acid replacements at the M4 transmembrane segment of the torpedo californica nicotinic acetylcholine receptor dramatically reduce the inhibition by 1-Propanol. Biophysical Soc. Abstr.
• Cruz-Martín, A., L.V. Rojas, J.L. Mercado, M.G. McNamee, and J.A. Lasalde-Dominicci. 2000. Tryptophan substitutions in lipid-exposed positions of the M3 transmembrane domains of the torpedo californica nicotinic acetylcholine receptor alter ion channel kinetics. Biophysical Soc. Abstr
• Mercado, J.L., A. Cruz-Martín, L.V. Rojas, M. McNamee, and J.A. Lasalde-Dominicci. 2000. Hydrophobic amino acid replacement at lipid exposed positions of the torpedo californica nicotinic acetylcholine receptor dramatically reduce the inhibition by propanol. FASEB Abstr..
• Cruz-Martín, A., J.L. Mercado, L.V. Rojas, M.G. McNamee, and J.A. Lasalde-Dominicci. 1999. Tryptophan substitutions in lipid-exposed positions of the gamma M3 and M4 transmembrane domains of the torpedo californica nicotinic acetylcholine receptor alter ion channel kinetics. Soc. Neurosci. Abstr.
• Cruz-Martín, A., J.L. Mercado, L.V. Rojas, M.G. McNamee, and J.A. Lasalde-Dominicci. 1999. Tryptophan substitutions at lipid-exposed positions of the gamma M3 transmembrane domain increase the macroscopic ionic current response of the torpedo californica nicotinic acetylcholine receptor. MBRS Abstr.