Ammonia affects the activity and expression of soluble and particulate GC in cultured rat astrocytes.
Neurotoxic effects of ammonia are mediated by increased accumulation of nitric oxide (NO), which combines with free radicals to form a highly toxic compound, peroxynitrite. Previous experiments in vivo and in vitro have suggested that this phenomenon engages neuron-derived NO and is coupled to changes in the accumulation of cGMP. The present study accounted for the facts that: (i) astrocytes, not neurons are the morphological target of ammonia, and (ii) both NO-dependent, soluble (sGC) and NO-independent, particulate guanylate cyclase (pGC) mediate cGMP production in the cells. Neocortical rat astrocytes were treated for 1 or 24 h with 5 mM ammonium chloride ("ammonia") and then subjected to: (i) cGMP measurement, and (ii) mRNA and/or protein expression analysis of alpha1 and beta1 subunits of sGC and two pGC forms: pGC-A and pGC-B. Treatment with ammonia for 1h increased accumulation of cGMP and sGCbeta1 mRNA expression, without producing significant changes in the protein expression. This was followed by a decrease of cGMP level at 24 h treatment, associated with a decreased expression of sGCbeta1 and sGCalpha1 mRNA and sGCbeta1 protein. Expression of pGC-A and pGC-B mRNA was elevated in ammonia-treated astrocytes after 24 h. Accordingly, increased cGMP accumulation was noted in the presence of a specific sGC inhibitor (ODQ). The results show that ammonia affects cGMP production in astrocytes, and that this may involve not only sGC but also pGC.
Konopacka
Agnieszka
A
Department of Neurotoxicology, Medical Research Centre, Polish Academy of Sciences, 02-106 Warsaw, Pawińskiego St. 5, Poland.
Freśko
Inez
I
Piaskowski
Sylwester
S
Albrecht
Jan
J
Zielińska
Magdalena
M
D000208:Acute Disease; D000641:Ammonia / Q000378:metabolism*; D000643:Ammonium Chloride / Q000633:toxicity; D000818:Animals; D001253:Astrocytes / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology; D006152:Cyclic GMP / Q000096:biosynthesis; D006162:Guanylate Cyclase / Q000096:biosynthesis* / Q000235:genetics; D022124:Hyperammonemia / Q000201:enzymology / Q000378:metabolism; D009474:Neurons / Q000201:enzymology / Q000378:metabolism; D021122:Protein Subunits / Q000096:biosynthesis / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D017208:Rats, Wistar; D018160:Receptors, Cytoplasmic and Nuclear / Q000096:biosynthesis* / Q000235:genetics; D000071756:Soluble Guanylyl Cyclase
D021122:Protein Subunits; D012333:RNA, Messenger; D018160:Receptors, Cytoplasmic and Nuclear; D000643:Ammonium Chloride; D000641:Ammonia; D006162:Guanylate Cyclase; D000071756:Soluble Guanylyl Cyclase; D006152:Cyclic GMP