Distribution of the P2X2 receptor and chemical coding in ileal enteric neurons of obese male mice(ob/ob)

AIM:To investigate the colocalization,density and profile of neuronal areas of enteric neurons in the ileum of male obese mice.METHODS:The small intestinal samples of male mice in an obese group(OG)(C57BL/6J ob/ob)and a control group(CG)(+/+)were used.The tissues were analyzed using a double immunostaining technique for immunoreactivity(ir)of the P2X2 receptor,nitric oxide synthase(NOS),choline acetyl transferase(ChAT)and calretinin(Calr).Also,we investigated the density and profile of neuronal areas of the NOS-,ChAT-and Calrir neurons in the myenteric plexus.Myenteric neurons were labeled using an NADH-diaphorase histochemical staining method.RESULTS:The analysis demonstrated that the P2X2receptor was expressed in the cytoplasm and in the nuclear and cytoplasmic membranes only in the CG.Neuronal density values(neuron/cm2)decreased 31%(CG:6579±837;OG:4556±407)and 16.5%(CG:7796±528;OG:6513±610)in the NOS-ir and calretininir neurons in the OG,respectively(P<0.05).Density of ChAT-ir(CG:6200±310;OG:8125±749)neurons significantly increased 31%in the OG(P<0.05).Neuron size studies demonstrated that NOS,ChAT,and Calr-ir neurons did not differ significantly between the CG and OG groups.The examination of NADH-diaphorase-positive myenteric neurons revealed an overall similarity between the OG and CG.CONCLUSION:Obesity may exert its effects by promoting a decrease in P2X2 receptor expression and modifications in the density of the NOS-ir,ChAT-ir and CalR-ir myenteric neurons.

Regulation of neural stem/progenitor cell functions by P2X and P2Y receptors

Neural stem/progenitor cells：Radial glial cells constitute multipotent cells in the ventricular zone,lining the wall of the lateral ventricle of the embryonic brain.They have the capacity to give rise to cells belonging to all three major linages（neurons,astrocytes and oligodendrocytes）of the nervous system（Tang and Illes,2017）.

Expression of the P2X_2 receptor in different classes of ileum myenteric neurons in the female obese ob/ob mouse

AIM:To examine whether the ob/ob mouse model of obesity is accompanied by enteric nervous system ab-normalities such as altered motility METHODS:The study examined the distribution of the P2X 2 receptor (P2X 2 R) in myenteric neurons of female ob/ob mice. Specifically, we used immunohistochemistry to analyze the co-expression of the P2X 2 R with neuronal nitric oxide synthase (nNOS), choline acetyltrans-ferase (ChAT), and calretinin (CalR) in neurons of the small intestine myenteric plexus in ob/ob and control female mice In these sections, we used scanning confocal microscopy to analyze the co-localization of these markers as well as the neuronal density (cm 2 ) and area profile (μm2) of P2X 2 R-positive neurons In addition, enteric neurons were labeled using the nicotinamide adenine dinucleotide (NA H) diaphorase method and analyzed with light microscopy as an alternate means by which to analyze neuronal density and areaRESULTS:In the present study, we observed a 29 6% increase in the body weight of the ob/ob animals (OG) compared to the control group (CG) In addition, the average small intestine area was increased by approxi-mately 29 6% in the OG compared to the CG Immu-noreactivity (IR) for the P2X 2 R, nNOS, ChAT and CalR was detectable in the myenteric plexus, as well as in the smooth muscle, in both groups This IR appeared to be mainly cytoplasmic and was also associated with the cell membrane of the myenteric plexus neurons, where it outlined the neuronal cell bodies and their processes P2X 2 R-IR was observed to co-localize 100% with that for nNOS, ChAT and CalR in neurons of both groups In the ob/ob group, however, we observed that the neuronal density (neuron/cm 2 ) of P2X 2 R-IR cells was in-creased by 62% compared to CG, while that of NOS-IR and ChAT-IR neurons was reduced by 49% and 57%, respectively, compared to control mice The neuronal density of CalR-IR neurons was not different between the groups Morphometric studies further demonstrated that the cell body profile area (μm2) of nNOS-IR, ChAT-IR and CalR-IR neurons was increased by 34%, 20% and 55%, respectively, in the OG compared to controls Staining for NA H diaphorase activity is widely used to detect alterations in the enteric nervous system; however, our qualitative examination of NA H-diaphorase positive neurons in the myenteric ganglia revealed an overall similarity between the two groups CONCLUSION:We demonstrate increases in P2X2R expression and alterations in nNOS, ChAT and CalR IR in ileal myenteric neurons of female ob/ob mice compared to wild-type controls.

Microencapsulation improves inhibitory effects of transplanted olfactory ensheathing cells on pain after sciatic nerve injury

Olfactory bulb tissue transplantation inhibits P2X2/3 receptor-mediated neuropathic pain. However, the olfactory bulb has a complex cellular composition, and the mechanism underlying the action of purified transplanted olfactory ensheathing cells（OECs） remains unclear. In the present study, we microencapsulated OECs in alginic acid, and transplanted free and microencapsulated OECs into the region surrounding the injured sciatic nerve in rat models of chronic constriction injury. We assessed mechanical nociception in the rat models 7 and 14 days after surgery by measuring paw withdrawal threshold, and examined P2X2/3 receptor expression in L4–5 dorsal root ganglia using immunohistochemistry. Rats that received free and microencapsulated OEC transplants showed greater withdrawal thresholds than untreated model rats, and weaker P2X2/3 receptor immunoreactivity in dorsal root ganglia. At 14 days, paw withdrawal threshold was much higher in the microencapsulated OEC-treated animals. Our results confirm that microencapsulated OEC transplantation suppresses P2X2/3 receptor expression in L4–5 dorsal root ganglia in rat models of neuropathic pain and reduces allodynia, and also suggest that transplantation of microencapsulated OECs is more effective than transplantation of free OECs for the treatment of neuropathic pain.

SpinalP2X7R contributes to streptozotocin-induced mechanical allodynia in mice

Painful diabetic neuropathy(PDN)is a diabetes mellitus complication.Unfortunately,the mechanisms underlying PDN are still poorly understood.Adenosine triphosphate(ATP)-gated P2X7 receptor(P2X7R)plays a pivotal role in non-diabetic neuropathic pain,but little is known about its effects on streptozotocin(STZ)-induced peripheral neuropathy.Here,we explored whether spinal cord P2X7R was correlated with the generation of mechanical allodynia(MA)in STZ-induced type 1 diabetic neuropathy in mice.MA was assessed by measuring paw withdrawal thresholds and western blotting.Immunohistochemistry was applied to analyze the protein expression levels and localization of P2X7R.STZ-induced mice expressed increased P2X7R in the dorsal horn of the lumbar spinal cord during MA.Mice injected intrathecally with a selective antagonist of P2X7R and P2X7R knockout(KO)mice both presented attenuated progression of MA.Double-immunofluorescent labeling demonstrated that P2X7R-positive cells were mostly co-expressed with Iba1(a microglia marker).Our results suggest that P2X7R plays an important role in the development of MA and could be used as a cellular target for treating PDN.