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Asian Citrus Psyllid (ACP), Diaphorina citri, is a key vector transmitting the causative agent of Huanglongbing (HLB) disease. Population growth of ACP is evident after feeding on plant flush shoots, as they only oviposit here. However, the underlying mechanism as to why flush shoots govern oviposition is unclear. This study compares the fecundity and ovarian morphology of ACP between young flush and mature leaves. Furthermore, the transcriptome of mated females infesting Murraya paniculata was analysed. Finally, the gene of the key Target of Rapamycin (TOR) signalling pathway was silenced by RNAi. Results indicated that flush shoot feeding activated the development of the psyllids ovary and therefore induced oviposition. A total of 126 and 2794 differentially expressed genes were detected at 1 and 5 days, respectively, after pest infestation of flush shoots compared to mature leaves. Many genes are involved in protein metabolism, Mitogen‐Activated Protein Kinase (MAPK) signalling pathway, hormone synthesis, and TOR signalling pathway: all thought to activate reproduction. Silencing of the positive regulator gene DcRheb in the TOR pathway resulted in lower levels of ecdysone and juvenile hormone and decreased vitellogenin synthesis, further disrupting reproductive ability. This study enhances understanding of the molecular mechanism underlying ACP's reproductive strategy. [ABSTRACT FROM AUTHOR]

Background We have previously shown that ingestion of Lactobacillus reuteri may modulate colonic enteric neuron activity but with unknown effects on colon motility. The aim of the present report was to elucidate the neuronal mechanisms of action of the probiotic by comparing the effects on motility of L. reuteri ingestion with blockade of a specific ionic current in enteric neurons. Methods We have used intraluminal pressure recordings from ex vivo rat colon segments and whole cell patch clamp recordings from neurons of rat longitudinal muscle myenteric plexus preparations to investigate the effects of L. reuteri and TRAM-34 on colon motility and neurophysiology. The effects of daily feeding of 109 L. reuteri bacteria or acute application of TRAM-34 on threshold fluid filling pressure or pulse pressure was measured. Key Results Lactobacillus reuteri increased intraluminal fluid filling pressure thresholds for evoking pressure pulses by 51% from 0.47 ± 0.17 hPa; the probiotic also decreased the pulse pressure amplitudes, but not frequency, by 18% from 3.91 ± 0.52 hPa. The intermediate conductance calcium-dependent potassium (IKCa) channel blocker TRAM-34 (3 μmol L−1) increased filling threshold pressure by 43% from 0.52 ± 0.22 hPa and reduced pulse pressure amplitude by 40% from 2.63 ± 1.11 hPa; contraction frequency was unaltered. TRAM-34 (3 μmol L−1) reduced membrane polarization, leak conductance and the slow afterhyperpolarization current in 16/16 myenteric rat colon AH cells but 19/19 S cells were unaffected. Conclusions & Inferences The present results are consistent with L. reuteri enhancing tonic inhibition of colon contractile activity by acting via the IKCa channel current in AH cells. [ABSTRACT FROM AUTHOR]

This case report presents a family with developmental glaucoma accompanied by microcornea resulting from novel mutations in the ADAMTS18 gene. The index case involves a 5-year-old twin brother, who, during a routine examination, exhibited elevated intraocular pressure persisting for over a month. The peak intraocular pressure reached approximately 25 mmHg (1 mmHg=0.133 kPa) in both eyes, with a corneal diameter of less than 10 mm. Ocular examination revealed an enlarged cup-to-disc ratio, and optical coherence tomography (OCT) demonstrated thinning of the retinal nerve fiber layer and ganglion cell layer. Ultrasound biomicroscopy combined with gonioscopy indicated partial angle closure and abnormal anterior chamber angle development. The ocular manifestations in the twin brother were consistent with those observed in the twin sister. The clinical diagnosis was bilateral developmental glaucoma with microcornea. Genetic sequencing identified two novel compound heterozygous mutations in the ADAMTS18 gene in the twins: Mutation 1 (M1) involving the variant site 1 (c.3436C>T:p.R1146W) and Mutation 2 (M2) involving the variant site 2 (c.1454T>G:p.F485C). Ocular examinations of four additional family members were normal. Genetic testing revealed that the twins' father and sister carried M1, while the index case's mother and brother carried M2. This report underscores a unique association between ADAMTS18 gene mutations and developmental glaucoma with microcornea within a familial context, emphasizing the importance of genetic screening for early diagnosis and targeted management strategies.

Nitric oxide synthase (NOS) requires the substrate L-arginine for NO production to support multiple gastrointestinal functions. We asked, 'Where do enzymes to regenerate L-arginine from L-citrulline exist?'. We examined loci of immunoreactivities in the canine gastrointestinal tract for arginosuccinate synthetase and arginosuccinate lyase, enzymes that resynthesize L-arginine from L-citrulline, in relation to the distribution of nNOS immunoreactivity or NADPH-diaphorase histochemistry. Arginosuccinate synthetase and lyase were present in many neurones and nerve fibres in the myenteric plexus of the lower oesophageal sphincter (LOS), antrum, pylorus, ileum and colon; in the submucosal plexus of ileum and colon; in longitudinal muscle of ileum and colon; and in nerve bundles in circular muscle everywhere. LOS muscle was also immunoreactive for both enzymes. Circular and longitudinal muscle cells of the ileum and colon and cells resembling interstitial cells of Cajal in the deep muscular plexus of the ileum and the submuscular plexus of the colon also appeared immunoreactive. In neurones, arginosuccinate synthetase and nNOS were usually co-localized. NADPH diaphorase activity was present in LOS and likely in pylorus, but not in muscularis externa of ileum or colon. We conclude that resynthesis of L-arginine probably occurs in enteric nerves, interstitial cells of Cajal (ICC) and LOS muscle; also apparently in some cells without NOS to utilize it.

Depolarization elicited outward K+ currents from canine lower esophageal sphincter (LES) muscle cells, primarily through iberiotoxin (IbTX)- and tetraethylammonium-sensitive Ca(2+)-dependent K+ channels. Current magnitudes varied with pipette Ca2+ concentration (EC50 = 108.5 nM). NG-nitro-L-arginine (L-NNA, 10(-4)M), IbTX (10(-8)M), or buffering intracellular Ca2+ to 8 nM decreased outward currents > 80%. Sodium nitroprusside (NaNP, 10(-4)M) restored L-NNA-inhibited or low intracellular Ca2+ concentration (not IbTX)-inhibited currents. L-NNA or IbTX application depolarized LES cells from -43 to -35 mV. NaNP restored the membrane potential to -46 mV after L-NNA but not after IbTX application. Nifedipine (30 microM) reduced outward currents and abolished or reduced L-NNA or NaNP effects, respectively. Immunocytochemistry revealed the presence of both argininosuccinate synthetase and argininosuccinate lyase in LES muscle cells. L-Citrulline, like L-arginine, reversed L-NNA inhibition of outward currents; only L-arginine reversed inhibition of outward currents by an antibody to argininosuccinate synthetase. Therefore, endogenous nitric oxide production, activated by Ca2+ entrance involving L-type Ca2+ channels, may continuously enhance outward currents to modulate LES muscle cell membrane potential and excitability.

1. Studies on canine lower oesophageal sphincter (LOS) evaluated the existence and function of a myogenic, nitric oxide synthase (NOS) by use of immunocytochemistry for NOS isozymes, NADPH-d histochemistry, [3H]-L-arginine to [3H]-L-citrulline transformation. In addition, functional studies in the muscle bath were performed. 2. Smooth muscle bundles or freshly isolated smooth muscle cells of LOS were NADPH-d reactive but did not recognize some antibodies against neural, endothelial or inducible NOS. NADPH-d reactivity and immunoreactivity to a neural NOS antibody were colocalized in LOS enteric nerves. Muscle plasma membrane-enriched fractions from fresh and cultured LOS cells converted [3H]-L-arginine to [3H]-L-citrulline; activity was mostly Ca2+/calmodulin-dependent. 3. N-Nitro-L-arginine (L-NOARG) persistently increased tone (blocked by L-arginine) in muscle strips despite blockade of nerve function. Nifedipine prevented or abolished L-NOARG-induced, but not carbachol-induced, contraction showing that tone increase by L-NOARG required functional L-Ca channels. 4. Membrane-bound, myogenic NOS in canine LOS may release NO continuously when Ca2+ entry through L-Ca channels occurs under physiological conditions and thereby modulate tone in LOS.

Our previous study demonstrated that PYY was a major neuropeptide in the canine enteric nervous system, the present study defines the locations of NPY-containing enteric neurons. NPY-positive nerve cell bodies and fibers were numerous in gastric and pyloric myenteric plexuses as were positive nerve fibers in antral and pyloric muscle layers, pyloric sphincter muscle layers, and surrounding blood vessels. In contrast to findings for PYY, there were considerably fewer NPY-positive nerve cell bodies and fibers in the canine ileum and colon. Noradrenaline was the predominant catecholamine in all plexuses, the rank order of its contents being: deep muscular > submucous > myenteric plexus. The dopamine/noradrenaline ratio was constant in all plexuses; adrenaline was present in minor amounts. PYY is the more abundant neuropeptide in ileum and colon, and NPY has a greater presence in the gastric antrum, pylorus, and surrounding blood vessels. PYY and NPY may play different functional roles in the GI tract.

Background: Vasoactive intestinal polypeptide (VIP) may be a nonadrenergic, noncholinergic inhibitory transmitter in the lower esophageal sphincter (LES). There is no biochemical evidence of VIP receptors in the LES., Methods: Using membranes from canine LES, VIP receptor distribution and characterization were analyzed by radioligand binding and cross-linking experiments., Results: High densities of saturable VIP receptors were found (maximum bound [Bmax], 539.2 fmol/mg in the synaptosome-enriched fraction [P2] and 732.7 fmol/mg in the smooth muscle, plasma membrane-enriched fraction [Mic II]), with high affinity for 125I-VIP (dissociation constant [Kd], 1.38 nmol/L in P2 and 1.40 nmol/L in Mic II). Competition binding studies suggested the presence of two binding sites, a high-affinity (inhibitor constant [Ki1], 0.064 nmol/L) and a low-affinity (Ki2, 2.68 nmol/L) binding site in P2 membranes, but only one binding site (Ki, 1.18 nmol/L) in Mic II membranes. Guanosine triphosphate-gamma-s pretreatment eliminated high-affinity binding in P2 membranes by conversion to binding sites of lower affinity (Ki, 2.82 nmol/L). Studies with a cross-linking agent identified VIP receptors in synaptosomal and smooth muscle plasma membrane fractions; a single polypeptide of approximately 60 kilodaltons was found in each membrane., Conclusions: Specific VIP receptors exist in both synaptosomal and smooth muscle plasma membrane of canine LES.

We have established the peptidase content of a P2 fraction (enriched in synaptosomes) and plasma membranes prepared from canine intestinal mucosa. Fourteen exo- and endopeptidases were assayed with fluorimetric or chromogenic substrates and identified by means of specific peptidase inhibitors. Post-proline dipeptidyl aminopeptidase IV, aminopeptidase M, and carboxypeptidase A were the most abundant exopeptidases, while aminopeptidases A and B, dipeptidyl aminopeptidase, pyroglutamyl peptide hydrolase I, and carboxypeptidase B displayed little, if any, activity. Endopeptidase 24.11 was the only endopeptidase that was detected in high amount. By contrast, proline endopeptidase exhibited a low activity, while angiotensin-converting enzyme, endopeptidase 24.15, endopeptidase 24.16, and cathepsin B and D-like activities were not detected. The catabolic rates of the two related neuropeptides, neurotensin (NT) and neuromedin N (NN), established that NN was inactivated 16 to 24 times faster than NT by plasma membrane and P2 fractions, respectively. Furthermore, the two peptides underwent qualitatively distinct mechanisms of degradation. A phosphoramidon-sensitive formation of NT(1-10) was detected as the major NT catabolite, indicating that NT was susceptible to an endoproteolytic cleavage elicited by endopeptidase 24.11. By contrast, NN was inactivated by the action of an exopeptidase at its N-terminus, leading to the formation of [des-Lys1]NN. The occurrence of this NN metabolite was prevented by bestatin and actinonin, but not by the aminopeptidase B inhibitor, arphamenine B, indicating that the release of the N-terminal residue of NN was likely due to aminopeptidase M.(ABSTRACT TRUNCATED AT 250 WORDS)

Bovine endothelial cells cultured from pulmonary artery (ATCC cell line No. 209) were found to contain a high density of 125I-VIP (vasoactive intestinal polypeptide) binding sites. These were found to be saturable and to be fit by a single binding site model (Kd 1.8 nM; Bmax 534 fmol/mg protein). Studies of association and dissociation of 125I-VIP to this site revealed that binding was fully reversible and yielded a Kd value similar to that from equilibrium binding. However competition studies showed that VIP competed for binding at two sites (Ki1 1.2 x 10(-11) M, Ki2 4.7 x 10(-9) M; N1 = 21%, N2 = 77%; Ki a dissociation constant for inhibitor; N percentage of occupied receptors). [Phe1]VIP also competed at two sites, but VIP-(10-28), PHM, [4-Cl-D-Phe6,Leu17]VIP and [D-Ala4]VIP displaced all specific VIP binding in a simple competitive manner. These VIP binding sites were shown to be functional. In patch clamp studies VIP 10(-8)-10(-7) M inhibited opening of inwardly rectifying K+ channels on hyperpolarization. These channels were affected appropriately by alteration in the K(+)-gradient and by Ba2+ or Cs+. The VIP antagonist [4-Cl-D-Phe6, Leu17]VIP prevented or reversed the effects of VIP. These results show that functional VIP receptors are present in high density in a endothelial cell line and provide a possible model for analysis of the molecular biology of these receptors.