title: L-citrulline inhibits [3H]acetylcholine release from rat motor nerve terminals by increasing adenosine outflow and activation of A1 receptors.
abstract: OBJECTIVE
  Nitric oxide (NO) production and depression of neuromuscular transmission are closely related, but little is known about the role of L-citrulline, a co-product of NO biosynthesis, on neurotransmitter release.
  
  
  METHODS
  Muscle tension recordings and outflow experiments were performed on rat phrenic nerve-hemidiaphragm preparations stimulated electrically.
  
  
  RESULTS
  L-citrulline concentration-dependently inhibited evoked [(3)H]ACh release from motor nerve terminals and depressed nerve-evoked muscle contractions. The NO synthase (NOS) substrate, L-arginine, and the NO donor, 3-morpholinosydnonimine chloride (SIN-1), also inhibited [(3)H]ACh release with a potency order of SIN-1>L-arginine>L-citrulline. Co-application of L-citrulline and SIN-1 caused additive effects. NOS inactivation with N(omega)-nitro-L-arginine prevented L-arginine inhibition, but not that of L-citrulline. The NO scavenger, haemoglobin, abolished inhibition of [(3)H]ACh release caused by SIN-1, but not that caused by L-arginine. Inactivation of guanylyl cyclase with 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) fully blocked SIN-1 inhibition, but only partially attenuated the effects of L-arginine. Reduction of extracellular adenosine accumulation with adenosine deaminase or with the nucleoside transport inhibitor, S-(p-nitrobenzyl)-6-thioinosine, attenuated the effects of L-arginine and L-citrulline, while not affecting inhibition by SIN-1. Similar results were obtained with the selective adenosine A(1) receptor antagonist, 1,3-dipropyl-8-cyclopentylxanthine. L-citrulline increased the resting extracellular concentration of adenosine, without changing that of the adenine nucleotides.
  
  
  CONCLUSIONS
  NOS catalyses the formation of two neuronally active products, NO and L-citrulline. While, NO may directly reduce transmitter release through stimulation of soluble guanylyl cyclase, the inhibitory action of L-citrulline may be indirect through increasing adenosine outflow and subsequently activating inhibitory A(1) receptors.
authors.affiliation: Laboratório de Farmacologia e Neurobiologia/UMIB, Instituto de Ciências Biomédicas de Abel Salazar, Universidade do Porto, Porto, Portugal.
authors.forename: A
authors.initials: A
authors.lastname: Barroso
authors.forename: L
authors.initials: L
authors.lastname: Oliveira
authors.forename: E
authors.initials: E
authors.lastname: Campesatto-Mella
authors.forename: C
authors.initials: C
authors.lastname: Silva
authors.forename: M A
authors.initials: MA
authors.lastname: Timóteo
authors.forename: M T
authors.initials: MT
authors.lastname: Magalhães-Cardoso
authors.forename: W
authors.initials: W
authors.lastname: Alves-do-Prado
authors.forename: P
authors.initials: P
authors.lastname: Correia-de-Sá
mesh_terms: D000109:Acetylcholine / Q000378:metabolism*; D000241:Adenosine / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001120:Arginine / Q000494:pharmacology; D002956:Citrulline / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D005260:Female; D008297:Male; D009046:Motor Neurons / Q000187:drug effects* / Q000378:metabolism; D009119:Muscle Contraction / Q000187:drug effects; D019001:Nitric Oxide Synthase / Q000502:physiology; D019335:Nitroarginine / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D043682:Receptor, Adenosine A1 / Q000187:drug effects* / Q000502:physiology
chemical_list: D043682:Receptor, Adenosine A1; D019335:Nitroarginine; D002956:Citrulline; D000255:Adenosine Triphosphate; D001120:Arginine; D019001:Nitric Oxide Synthase; D000241:Adenosine; D000109:Acetylcholine