Photochromic ligands have been used to control a variety of biological functions, especially in neural systems. Recently, much effort has been invested in the photocontrol of ion channels and G-protein coupled receptors found in the synapse. Herein, we describe the expansion of our photopharmacological approach toward the remote control of an enzyme. Building on hallmark studies dating from the late 1960s, we evaluated photochromic inhibitors of one of the most important enzymes in synaptic transmission, acetylcholinesterase (AChE). Using structure-based design, we synthesized several azobenzene analogues of the well-known AChE inhibitor tacrine (THA) and determined their effects on enzymatic activity. One of our compounds, AzoTHA, is a reversible photochromic blocker of AChE in vitro and ex vivo with high affinity and fast kinetics. As such, AzoTHA can be used to control synaptic transmission on the neuromuscular endplate based on the light-dependent clearance of a neurotransmitter.
Title: Photoregulation of biological activity by photocromic reagents. II. Inhibitors of acetylcholinesterase Bieth J, Vratsanos SM, Wassermann N, Erlanger BF Ref: Proc Natl Acad Sci U S A, 64:1103, 1969 : PubMed
The enzymic activity of acetylcholinesterase can be photoregulated through the mediation of photochromic inhibitors of the enzyme. N-p-phenylazophenyl-N-phenylcarbamyl fluoride, an irreversible inhibitor of acetylcholinesterase, exists as two geometric isomers which are interconvertible through the action of light. The cis isomer, which predominates after exposure to light of 320 nm, is more active than the trans isomer, which results from exposure to light of 420 nm. It was possible, therefore, to use light energy to regulate the inactivation of the enzyme. Similarly, levels of acetylcholinesterase activity could be photo-regulated in a completely reversible manner by means of the photochromic reversible inhibitor p-phenylazophenyltrimethylammonium chloride. These experiments can serve as models for similar phenomena observed in nature, particularly in photoperiodic rhythms of higher animals.
Title: Photoregulation of biological activity by photochromic reagents. 3. Photoregulation of bioelectricity by acetylcholine receptor inhibitors Deal WJ, Erlanger BF, Nachmansohn D Ref: Proc Natl Acad Sci U S A, 64:1230, 1969 : PubMed
The photochromic compounds N-p-phenylazophenyl-N-phenylcarbamylcholine chloride and p-phenylazophenyltrimethylammonium chloride inhibit the carbamylcholine-produced depolarization of the excitable membrane of the monocellular electroplax preparation of Electrophorus. The trans isomer of each predominates in the light of a photoflood (420 mmu) lamp; they are stronger inhibitors than the cis isomers, which predominate under ultraviolet (320 mmu) irradiation. The potential difference across the excitable membrane may be photoregulated by exposing an electroplax in the presence of a solution of carbamylcholine and either of the two compounds to light of appropriate wavelengths, since light shifts the cis-trans equilibrium. The system may be considered as a model illustrating how one may link a cis-trans isomerization, the first step in the initiation of a visual impulse, with substantial changes (20-30 mv) in the potential difference across an excitable membrane.
