Effects of protein deprivation and re-feeding on P2X2 receptors in enteric neurons.
OBJECTIVE
To investigate the effects of malnutrition and re-feeding on the P2X(2) receptor, nitric oxide synthase (NOS), calretinin, calbindin and choline acetyltransferase (ChAT) in neurons of the rat ileum.


METHODS
We analyzed the co-localization, numbers and sizes of P2X(2)-expressing neurons in relation to NOS-immunoreactive (IR), calbindin-IR, ChAT-IR, and calretinin-IR neurons of the myenteric and submucosal plexus. The experimental groups consisted of: (1) rats maintained on normal feed throughout pregnancy until 42 d post-parturition (N); (2) rats deprived of protein throughout pregnancy and 42 d post-parturition (D); and (3) rats undernourished for 21 d post-parturition and then given a protein diet from days 22 to 42 (DR). The myenteric and submucosal plexuses were evaluated by double labeling by immunohistochemical methods for P2X(2) receptor, NOS, ChAT, calbindin and calretinin.


RESULTS
We found similar P2X(2) receptor immunoreactivity in the cytoplasm and surface membranes of myenteric and submucosal neurons from the N, D and DR groups. Double labeling of the myenteric plexus demonstrated that approximately 100% of NOS-IR, calbindin-IR, calretinin-IR and ChAT-IR neurons in all groups also expressed the P2X(2) receptor. In the submucosal plexus, the calretinin-IR, ChAT-IR and calbindin-IR neurons were nearly all immunoreactive for the P2X(2) receptor. In the myenteric plexus, there was a 19% increase in numbers per cm(2) for P2X(2) receptor-IR neurons, 64% for NOS-IR, 84% for calretinin-IR and 26% for ChAT-IR neurons in the D group. The spatial density of calbindin-IR neurons, however, did not differ among the three groups. The submucosal neuronal density increased for calbindin-IR, calretinin-IR and ChAT-IR neurons. The average size of neurons in the myenteric plexus neurons in the D group was less than that in the controls and, in the re-fed rats; there was a 34% reduction in size only for the calretinin-IR neurons.


CONCLUSIONS
This work demonstrates that expression of the P2X(2) receptor is present in inhibitory, intrinsic primary afferent, cholinergic secretomotor and vasomotor neurons. Undernutrition affected P2X(2) receptor expression in the submucosal plexus, and neuronal and size. These changes were rescued in the re-fed rats.
Misawa
Rúbia
R
Department of Anatomy, Institute of Biomedical Sciences, University of São Paulo, Av. Prof. Dr. Lineu Prestes 2415, São Paulo, Brazil.
Girotti
Priscila Azevedo
PA
Mizuno
Márcia Sanae
MS
Liberti
Edson Aparecido
EA
Furness
John Barton
JB
Castelucci
Patricia
P
D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D064032:Calbindin 2; D064026:Calbindins; D002795:Choline O-Acetyltransferase / Q000378:metabolism; D004044:Dietary Proteins; D004435:Eating / Q000502:physiology*; D017615:Enteric Nervous System / Q000166:cytology*; D005260:Female; D007082:Ileum / Q000166:cytology / Q000294:innervation / Q000378:metabolism; D008297:Male; D009474:Neurons / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000378:metabolism; D011247:Pregnancy; D011488:Protein Deficiency / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D018048:Receptors, Purinergic P2 / Q000378:metabolism*; D058476:Receptors, Purinergic P2X2; D064030:S100 Calcium Binding Protein G / Q000378:metabolism
D015415:Biomarkers; C576512:Calb2 protein, rat; D064032:Calbindin 2; D064026:Calbindins; D004044:Dietary Proteins; C546403:P2rx2 protein, rat; D018048:Receptors, Purinergic P2; D058476:Receptors, Purinergic P2X2; D064030:S100 Calcium Binding Protein G; D019001:Nitric Oxide Synthase; D002795:Choline O-Acetyltransferase