SABER UCV >
1) Investigación >
Artículos Publicados >
Por favor, use este identificador para citar o enlazar este ítem:
http://hdl.handle.net/10872/13237
|
Título : | A novel PDE1A coupled to M2AChR at plasma membranes from bovine tracheal smooth muscle |
Autor : | Mastromatteo-Alberga, Patrizzia Placeres-Uray, Fabiola A Alfonzo-González, Marcelo Gonzalez de Alfonzo, Ramona Lippo de Becemberg, Itala Alfonzo R., Marcelo J. |
Palabras clave : | Airway smooth muscle asthma m2AChR muscarinic antagonists |
Fecha de publicación : | 2-Nov-2015 |
Editorial : | Journal of Receptors and Signal Transduction |
Resumen : | Muscarinic antagonists, via muscarinic receptors increase the cAMP/cGMP levels at bovine
tracheal smooth muscle (BTSM) through the inhibition of phosphodiesterases (PDEs), displaying
a similar behavior of vinpocetine (a specific-PDE1 inhibitor). The presence of PDE1 hydrolyzing
both cyclic nucleotides in BTSM strips was revealed. Moreover, a vinpocetine and muscarinic
antagonists inhibited PDE1 located at plasma membranes (PM) fractions from BTSM showing
such inhibition, an M2AChR pharmacological profile. Therefore, a novel Ca2+/CaM dependent
and vinpocetine inhibited PDE1 was purified and characterized at PM fractions from BTSM. This
PDE1 activity was removed from PM fractions using a hypotonic buffer and purified some 38
fold using two columns (Q-Sepharose and CaM-agarose). This PDE1 was stimulated by CaM and
inhibited by vinpocetine showing two bands in PAGE-SDS (56, 58 kDa) being the 58 kDa
identified as PDE1A by Western blotts. This PDE1A activity was assayed with [3H]cGMP and
[3H]cAMP exhibiting a higher affinity as Km (mM) for cGMP than cAMP but being close values
with Vmax cAMP/cGMP ratio of 1.5. The co-factor Mg2+ showed similar K(A) (mM) for both cyclic
nucleotides. Vinpocetine showed similar inhibition concentration 50% (IC50 of 4.9 and 4.6 mM)
for cAMP and cGMP, respectively. CaM stimulated the cyclic nucleotides hydrolysis by PDE1A
exhibiting similar activation constant as K(CaM), in nM range. The original finding was the
identification and purification of a vinpocetine and muscarinic antagonist-inhibited and CaMactivated
PM-bound PDE1A, linked to M2AChR. A model of this novel signal transducing
cascade for the regulation of cyclic nucleotides levels at BTSM is proposed.
Abbreviations: ASM: Airway smooth muscle; BTSM: Bovine Tracheal Smooth Muscle;
Benzamidine: amidinobenzene hydrochloride; CaM: calmodulin; CCh: carbamylcholine;
cAMP: cyclic adenosine 30, 50-monophosphate; cGMP: cyclic guanosine 30,50- monophosphate;
4-DAMP: 4-diphenylacetoxy-N-methylpiperidine; DTT: 1:4-dithiothreitol; PDE1A: phosphodiesterase
1A; PDE1B: phosphodiesterase tipo 1B; PDE1C: phosphodiesterase tipo 1C; PMSF:
phenylmethylsulphonylfluoride; IBMX: isobutyl-methylxanthine; GPCR: G protein-coupled
receptors; mAchR or MAchR: muscarinic acetylcholine receptor; M2AChR: M2 muscarinic
receptor; M3AChR: M3 muscarinic receptor; NO-sGC: NO sensitive soluble guanylyl cyclase;
NPR-GC: Natriuretic Peptides Receptor guanylyl cyclase; TCA: Trichloroacetic acid; Vinpocetine:
(3a: 16b)-eburnamenine-14-carboxylic ethyl ester acid |
URI : | http://hdl.handle.net/10872/13237 |
ISSN : | 1079-9893 |
Aparece en las colecciones: | Artículos Publicados
|
Los ítems de DSpace están protegidos por copyright, con todos los derechos reservados, a menos que se indique lo contrario.
|