ABSTRACT
1. Previous studies have shown that phorbol esters induce protein kinase C (PKC) mediated phosphorylation of the vesicular acetylcholine transporter (VAChT) and change its interaction with vesamicol. However, it is not clear whether physiological activation of receptors coupled to PKC activation can alter VAChT behavior. 2. Here we tested whether activation of kaianate (KA) receptors alters VAChT. Several studies suggest that the cholinergic amacrine cells display KA/AMPA receptors that mediate excitatory input to these neurons. In addition, KA in the chicken retina can generate intracellular messengers with the potential to regulate cellular functions. 3. In cultured chicken retina (E8C11) KA reduced vesamicol binding to VAChT by 53%. This effect was potentiated by okadaic acid, a protein phosphatase inhibitor, and was totally prevented by BIM, a PKC inhibitor. 4. Phorbol myristate acetate (PMA), but not alpha-PMA, reduced in more than 85% the number of L-[3H]-vesamicol-specific binding sites in chicken retina, confirming that activation of PKC can influence vesamicol binding to chicken VAChT. 5. The data show that activation of glutamatergic receptors reduces [3H]-vesamicol binding sites (VAChT) likely by activating PKC and increasing the phosphorylation of the ACh carrier.
Subject(s)
Acetylcholine/metabolism , Carrier Proteins/metabolism , Membrane Transport Proteins , Presynaptic Terminals/metabolism , Protein Kinase C/metabolism , Receptors, Kainic Acid/metabolism , Retina/metabolism , Synaptic Transmission/physiology , Vesicular Transport Proteins , Animals , Binding Sites/drug effects , Binding Sites/physiology , Chick Embryo , Choline O-Acetyltransferase/metabolism , Enzyme Inhibitors/pharmacology , Immunohistochemistry , Kainic Acid/pharmacology , Organ Culture Techniques , Phosphorylation/drug effects , Piperidines/pharmacology , Presynaptic Terminals/drug effects , Protein Kinase C/antagonists & inhibitors , Receptors, Kainic Acid/drug effects , Retina/drug effects , Synaptic Transmission/drug effects , Tetradecanoylphorbol Acetate/pharmacology , Up-Regulation/drug effects , Up-Regulation/physiology , Vesicular Acetylcholine Transport ProteinsABSTRACT
Se reporta la existencia de un factor soluble endógeno de bajo peso molecular, derivado de la corteza frontal humana, el cual es modular la unión de [3H]-QNB al receptor colinérgico muscarínico. El efecto de esta sustancia fue de naturaleza no competitiva, ya que afectaba la capacidad máxima de unión, pero no la afinidad; sin embargo, un perfil bifásico, con inhibición hasta un 88-90 por ciento de la unión y una ulterior recuperación de hasta 50-60 por ciento de la inhibición, también fue observado. La acción de esta sustancia pareció ser reversible, resistente a la acción de la tripsina, termoestable, y presentó un peso molecular no mayor de 10.000 Da. Los resultados sugieren la posible existencia de heterogeneidad molecular en el factor endógeno aislado o la presencia de multiples sitios de acción sobre los cuales actuaría este factor
