The Protective Role of Intermedin in Contrast-Induced Acute Kidney Injury: Enhancing Peritubular Capillary Endothelial Cell Adhesion and Integrity Through the cAMP/Rac1 Pathway.

Contrast-induced acute kidney injury (CIAKI) is a common complication with limited treatments. Intermedin (IMD), a peptide belonging to the calcitonin gene-related peptide family, promotes vasodilation and endothelial stability, but its role in mitigating CIAKI remains unexplored. This study investigates the protective effects of IMD in CIAKI, focusing on its mechanisms, particularly the cAMP/Rac1 signaling pathway. Human umbilical vein endothelial cells (HUVECs) were treated with iohexol to simulate kidney injury in vitro. The protective effects of IMD were assessed using CCK8 assay, flow cytometry, ELISA, and Western blotting. A CIAKI rat model was utilized to evaluate renal peritubular capillary endothelial cell injury and renal function through histopathology, immunohistochemistry, immunofluorescence, Western blotting, and transmission electron microscopy. In vitro, IMD significantly enhanced HUVEC viability and mitigated iohexol-induced toxicity by preserving intercellular adhesion junctions and activating the cAMP/Rac1 pathway, with Rac1 inhibition attenuating these protective effects. In vivo, CIAKI caused severe damage to peritubular capillary endothelial cell junctions, impairing renal function. IMD treatment markedly improved renal function, an effect negated by Rac1 inhibition. IMD protects against renal injury in CIAKI by activating the cAMP/Rac1 pathway, preserving peritubular capillary endothelial integrity and alleviating acute renal injury from contrast media. These findings suggest that IMD has therapeutic potential in CIAKI and highlight the cAMP/Rac1 pathway as a promising target for preventing contrast-induced acute kidney injury in at-risk patients, ultimately improving clinical outcomes.

Allatotropin (AT) related peptides L-ATRP and D2-ATRP diastereomers activate an endogenous receptor and suppress heart rate in the Pacific abalone Haliotis discus hannai.

Allatotropin (AT) has been identified in many insects and plays important roles in the regulation of their intestinal contraction, heart rate, ion transport, and digestive enzyme secretion. However, information on AT-related bioinformatics in other animal phyla is scarce. In this study, we cloned a full-length cDNA encoding the AT-related peptide receptor (ATRPR) of the abalone Haliotis discus hannai (Hdh) and further characterized Hdh-ATRPR with its potential ligands, Hdh-ATRPs. In luciferase reporter and Ca2+ mobilization assays, Hdh-ATRPs, including a D-type Phe at the second amino acid position, Hdh-D2-ATRP, activated Hdh-ATRPR in a dose-dependent manner, whereas all-L-type Hdh-ATRP was a more potent ligand than Hdh-D2-ATRP. Furthermore, Hdh-ATRPs induced ERK1/2 phosphorylation in Hdh-ATRPR-expressing HEK293 cells, which was dose-dependently abolished by the PKC inhibitor Gö6983. The heart rate decreased significantly within 10 min when Hdh-D2-ATRP was injected into the adduct muscle sinus of abalone (0.2 or 1.0 µg/g body weight), while the abalone injected with a high concentration of Hdh-D2-ATRP (1.5 µg/g body weight) were sublethal within 5 h. Thus, Hdh-ATRP signaling is primarily linked to the Gαq/PKC and is possibly associated with heart rate regulation in abalone.

Isolation and characterization of NGFFYamide neuropeptide from Patiria pectinifera pyloric caeca extract.

Neuropeptides are small molecules that mediate intercellular signaling and regulate physiological processes. Starfish possess various myoactive neuropeptides, including starfish myorelaxant peptide (SMP) and a calcitonin-type peptide with apical muscle relaxing properties. In this study, we report the purification of a neuropeptide from starfish (Patiria pectinifera) pyloric caeca extract using high-performance liquid chromatography (HPLC) and an in vitro bioassay to screen for fractions and peptides with relaxing effects on P. pectinifera apical muscle preparations. A series of HPLC steps using reversed-phase and cation-exchange columns yielded a purified peptide with muscle-relaxing effects. The purified peptide's structure was determined by LC-MS and Edman degradation, revealing a pentapeptide with an amidated C-terminus (NGFFYamide) and a molecular mass of 646.2930 Da. This is the first report of NGFFYamide purification from P. pectinifera through biochemical methods. The nucleotide sequence encoding the NGFFYamide precursor was determined, showing the presence of a conserved neurophysin domain in the C-terminal region. RT-qPCR results confirmed high expression in radial nerves cord, consistent with previous findings on NG peptides in echinoderms. In vitro pharmacological studies on muscle preparations from P. pectinifera and Asterias amurensis revealed differential relaxing activity of NGFFYamide on apical muscles, while its effects on tube foot preparations were similar in both species. NGFFYamide also induced potent contraction in P. pectinifera cardiac stomach. Comparison of three NG peptides (NGFFYamide, NGFFFamide, and NGIWYamide) on P. pectinifera cardiac stomach revealed varying potency, suggesting class-specific receptor interactions. Tachyphylaxis was observed in P. pectinifera apical muscle but not in A. amurensis, warranting further investigation. Based on these results, it is plausible that NGFFYamide could be involved in regulating locomotion and feeding behavior in P. pectinifera, consistent with findings in Asterias rubens.

MP-13, a novel chimeric peptide of morphiceptin and pepcan-9, produces potent antinociception with limited side effects.

Pharmacological investigations have substantiated the potential of bifunctional opioid/cannabinoid agonists in delivering potent analgesia while minimizing adverse reactions. Peptide modulators of cannabinoid receptors, known as pepcans, have been investigated before. In this study, we designed a series of chimeric peptides based on pepcans and morphiceptin (YPFP-NH2). Here, we combined injections of pepcans and morphiceptin to investigate the combination treatment of opioids and cannabis and compared the analgesic effect with chimeric compounds. Subsequently, we employed computational docking to screen the compounds against opioid and cannabinoid receptors, along with an acute pain model, to identify the most promising peptide. Among these peptides, MP-13, a morphiceptin and pepcan-9 (PVNFKLLSH) construct, exhibited superior supraspinal analgesic efficacy in the tail-flick test, with an ED50 value at 1.43 nmol/mouse, outperforming its parent peptides and other chimeric analogs. Additionally, MP-13 displayed potent analgesic activity mediated by mu-opioid receptor (MOR), delta-opioid receptor (DOR), and cannabinoid type 1 (CB1) receptor pathways. Furthermore, MP-13 did not induce psychological dependence and gastrointestinal motility inhibition at the effective analgesic doses, and it maintained non-tolerance-forming antinociception throughout a 7-day treatment regimen, with an unaltered count of microglial cells in the periaqueductal gray region, supporting this observation. Moreover, intracerebroventricular administration of MP-13 demonstrated dose-dependent antinociception in murine models of neuropathic, inflammatory, and visceral pain. Our findings provide promising insights for the development of opioid/cannabinoid peptide agonists, addressing a crucial gap in the field and holding significant potential for future research and development. PERSPECTIVE: This article offers insights into the combination treatment of pepcans with morphiceptin. Among the chimeric peptides, MP-13 exhibited potent analgesic effects in a series of preclinical pain models with a favorable side-effect profile.

Neuropeptide Modulation Enables Biphasic Internetwork Coordination via a Dual-Network Neuron.

Linked rhythmic behaviors, such as respiration/locomotion or swallowing/chewing, often require coordination for proper function. Despite its prevalence, the cellular mechanisms controlling coordination of the underlying neural networks remain undetermined in most systems. We use the stomatogastric nervous system of the crab Cancer borealis to investigate mechanisms of internetwork coordination, due to its small, well-characterized feeding-related networks (gastric mill [chewing, ∼0.1 Hz]; pyloric [filtering food, ∼1 Hz]). Here, we investigate coordination between these networks during the Gly1-SIFamide neuropeptide modulatory state. Gly1-SIFamide activates a unique triphasic gastric mill rhythm in which the typically pyloric-only LPG neuron generates dual pyloric-plus gastric mill-timed oscillations. Additionally, the pyloric rhythm exhibits shorter cycles during gastric mill rhythm-timed LPG bursts, and longer cycles during IC, or IC plus LG gastric mill neuron bursts. Photoinactivation revealed that LPG is necessary to shorten pyloric cycle period, likely through its rectified electrical coupling to pyloric pacemaker neurons. Hyperpolarizing current injections demonstrated that although LG bursting enables IC bursts, only gastric mill rhythm bursts in IC are necessary to prolong the pyloric cycle period. Surprisingly, LPG photoinactivation also eliminated prolonged pyloric cycles, without changing IC firing frequency or gastric mill burst duration, suggesting that pyloric cycles are prolonged via IC synaptic inhibition of LPG, which indirectly slows the pyloric pacemakers via electrical coupling. Thus, the same dual-network neuron directly conveys excitation from its endogenous bursting and indirectly funnels synaptic inhibition to enable one network to alternately decrease and increase the cycle period of a related network.

A Novel Peptidomimetic Insecticide: Dippu-AstR-Based Rational Design and Biological Activity of Allatostatin Analogs.

Insect neuropeptides play an essential role in regulating growth, development, reproduction, nerve conduction, metabolism, and behavior in insects; therefore, G protein-coupled receptors of neuropeptides are considered important targets for designing green insecticides. Cockroach-type allatostatins (ASTs) (FGLamides allatostatins) are important insect neuropeptides in Diploptera punctata that inhibit juvenile hormone (JH) synthesis in the corpora allata and affect growth, development, and reproduction of insects. Therefore, the pursuit of novel insecticides targeting the allatostatin receptor (AstR) holds significant importance. Previously, we identified an AST analogue, H17, as a promising candidate for pest control. Herein, we first modeled the 3D structure of AstR in D. punctata (Dippu-AstR) and predicted the binding mode of H17 with Dippu-AstR to study the critical interactions and residues favorable to its bioactivity. Based on this binding mode, we designed and synthesized a series of H17 derivatives and assessed their insecticidal activity against D. punctata. Among them, compound Q6 showed higher insecticidal activity than H17 against D. punctata by inhibiting JH biosynthesis, indicating that Q6 is a potential candidate for a novel insect growth regulator (IGR)-based insecticide. Moreover, Q6 exhibited insecticidal activity against Plutella xylostella, indicating that these AST analogs may have a wider insecticidal spectrum. The underlying mechanisms and molecular conformations mediating the interactions of Q6 with Dippu-AstR were explored to understand its effects on the bioactivity. The present work clarifies how a target-based strategy facilitates the discovery of new peptide mimics with better bioactivity, enabling improved IGR-based insecticide potency in sustainable agriculture.

Intracerebroventricular PROK2 infusion could increase the secretion of male reproductive hormones by stimulating the HPG axis.

BACKGROUND: Prokineticin 2 (PROK2), an important neuropeptide that plays a key role in the neuronal migration of gonadotropin-releasing hormone (GnRH) in the hypothalamus, is known to have regulatory effects on the gonads. In the present study, the impact of intracerebroventricular (icv) PROK2 infusion on hypothalamic-pituitary-gonadal axis (HPG) hormones, testicular tissues, and sperm concentration was investigated. METHODS AND RESULTS: Rats were randomly divided into four groups: control, sham, PROK2 1.5 and PROK2 4.5. Rats in the PROK2 1.5 and PROK2 4.5 groups were administered 1.5 nmol and 4.5 nmol PROK2 intracerebroventricularly for 7 days via an osmotic mini pump (1 µl/h), respectively. Rat blood serum follicle stimulating hormone (FSH), luteinizing hormone (LH) and testosterone hormone levels were determined with the ELISA method in the blood samples after 7 days of infusion. GnRH mRNA expression was determined with the RT-PCR in hypothalamus tissues. analyze Sperm concentration was determined, and testicular tissue was examined histologically with the hematoxylin-eosin staining method. It was observed that GnRH mRNA expression increased in both PROK2 infusion groups. Serum FSH, LH and testosterone hormone levels also increased in these groups. Although sperm concentration increased in PROK2 infusion groups when compared to the control and sham, the differences were not statistically significant. Testicular tissue seminiferous epithelial thickness was higher in the PROK2 groups when compared to the control and sham groups. CONCLUSION: The present study findings demonstrated that icv PROK2 infusion induced the HPG axis. It could be suggested that PROK2 could be a potential agent in the treatment of male infertility induced by endocrinological defects.

Neuropeptide modulation of bidirectional internetwork synapses.

Oscillatory networks underlying rhythmic motor behaviors, and sensory and complex neural processing, are flexible, even in their neuronal composition. Neuromodulatory inputs enable neurons to switch participation between networks or participate in multiple networks simultaneously. Neuromodulation of internetwork synapses can both recruit and coordinate a switching neuron in a second network. We previously identified an example in which a neuron is recruited into dual-network activity via peptidergic modulation of intrinsic properties. We now ask whether the same neuropeptide also modulates internetwork synapses for internetwork coordination. The crab (Cancer borealis) stomatogastric nervous system contains two well-defined feeding-related networks (pyloric, food filtering, ∼1 Hz; gastric mill, food chewing, ∼0.1 Hz). The projection neuron MCN5 uses the neuropeptide Gly1-SIFamide to recruit the pyloric-only lateral posterior gastric (LPG) neuron into dual pyloric- plus gastric mill-timed bursting via modulation of LPG's intrinsic properties. Descending input is not required for a coordinated rhythm, thus intranetwork synapses between LPG and its second network must underlie coordination among these neurons. However, synapses between LPG and gastric mill neurons have not been documented. Using two-electrode voltage-clamp recordings, we found that graded synaptic currents between LPG and gastric mill neurons (lateral gastric, inferior cardiac, and dorsal gastric) were primarily negligible in saline, but were enhanced by Gly1-SIFamide. Furthermore, LPG and gastric mill neurons entrain each other during Gly1-SIFamide application, indicating bidirectional, functional connectivity. Thus, a neuropeptide mediates neuronal switching through parallel actions, modulating intrinsic properties for recruitment into a second network and as shown here, also modulating bidirectional internetwork synapses for coordination.NEW & NOTEWORTHY Neuromodulation can enable neurons to simultaneously coordinate with separate networks. Both recruitment into, and coordination with, a second network can occur via modulation of internetwork synapses. Alternatively, recruitment can occur via modulation of intrinsic ionic currents. We find that the same neuropeptide previously determined to modulate intrinsic currents also modulates bidirectional internetwork synapses that are typically ineffective. Thus, complementary modulatory peptide actions enable recruitment and coordination of a neuron into a second network.

Anorexigenic neuropeptides as anti-obesity and neuroprotective agents: exploring the neuroprotective effects of anorexigenic neuropeptides.

Since 1975, the incidence of obesity has increased to epidemic proportions, and the number of patients with obesity has quadrupled. Obesity is a major risk factor for developing other serious diseases, such as type 2 diabetes mellitus, hypertension, and cardiovascular diseases. Recent epidemiologic studies have defined obesity as a risk factor for the development of neurodegenerative diseases, such as Alzheimer's disease (AD) and other types of dementia. Despite all these serious comorbidities associated with obesity, there is still a lack of effective antiobesity treatment. Promising candidates for the treatment of obesity are anorexigenic neuropeptides, which are peptides produced by neurons in brain areas implicated in food intake regulation, such as the hypothalamus or the brainstem. These peptides efficiently reduce food intake and body weight. Moreover, because of the proven interconnection between obesity and the risk of developing AD, the potential neuroprotective effects of these two agents in animal models of neurodegeneration have been examined. The objective of this review was to explore anorexigenic neuropeptides produced and acting within the brain, emphasizing their potential not only for the treatment of obesity but also for the treatment of neurodegenerative disorders.

Long-term enhancements in antidepressant efficacy and neurogenesis: Effects of intranasal co-administration of neuropeptide Y 1 receptor (NPY1R) and galanin receptor 2 (GALR2) agonists in the ventral hippocampus.

This study evaluates the sustained antidepressant-like effects and neurogenic potential of a 3-day intranasal co-administration regimen of galanin receptor 2 (GALR2) agonist M1145 and neuropeptide Y Y1 receptor (NPY1R) agonist [Leu31, Pro34]NPY in the ventral hippocampus of adult rats, with outcomes analyzed 3 weeks post-treatment. Utilizing the forced swimming test (FST), we found that this co-administration significantly enhances antidepressant-like behaviors, an effect neutralized by the GALR2 antagonist M871, highlighting the synergistic potential of these neuropeptides in modulating mood-related behaviors. In situ proximity ligation assay (PLA) indicated a significant increase in GALR2/NPYY1R heteroreceptor complexes in the ventral hippocampal dentate gyrus, suggesting a molecular basis for the behavioral outcomes observed. Moreover, proliferating cell nuclear antigen (PCNA) immunolabeling revealed increased cell proliferation in the subgranular zone of the dentate gyrus, specifically in neuroblasts as evidenced by co-labeling with doublecortin (DCX), without affecting quiescent neural progenitors or astrocytes. The study also noted a significant uptick in the number of DCX-positive cells and alterations in dendritic morphology in the ventral hippocampus, indicative of enhanced neuronal differentiation and maturation. These morphological changes highlight the potential of these agonists to facilitate the functional integration of new neurons into existing neural circuits. By demonstrating the long-lasting effects of a brief, 3-day intranasal administration of GALR2 and NPY1R agonists, our findings contribute significantly to the understanding of neuropeptide-mediated neuroplasticity and herald novel therapeutic strategies for the treatment of depression and related mood disorders, emphasizing the therapeutic promise of targeting neurogenesis and neuronal maturation processes.

Orexins and primary headaches: an overview of the neurobiology and clinical impact.

INTRODUCTION: Primary headaches, including migraines and cluster headaches, are highly prevalent disorders that significantly impact quality of life. Several factors suggest a key role for the hypothalamus, including neuroimaging studies, attack periodicity, and the presence of altered homeostatic regulation. The orexins are two neuropeptides synthesized almost exclusively in the lateral hypothalamus with widespread projections across the central nervous system. They are involved in an array of functions including homeostatic regulation and nociception, suggesting a potential role in primary headaches. AREAS COVERED: This review summarizes current knowledge of the neurobiology of orexins, their involvement in sleep-wake regulation, nociception, and functions relevant to the associated symptomology of headache disorders. Preclinical reports of the antinociceptive effects of orexin-A in preclinical models are discussed, as well as clinical evidence for the potential involvement of the orexinergic system in headache. EXPERT OPINION: Several lines of evidence support the targeted modulation of orexinergic signaling in primary headaches. Critically, orexins A and B, acting differentially via the orexin 1 and 2 receptors, respectively, demonstrate differential effects on trigeminal pain processing, indicating why dual-receptor antagonists failed to show clinical efficacy. The authors propose that orexin 1 receptor agonists or positive allosteric modulators should be the focus of future research.

Discovery of Nivasorexant (ACT-539313): The First Selective Orexin-1 Receptor Antagonist (SO1RA) Investigated in Clinical Trials.

The orexin system consists of two neuropeptides (orexins A and B) and two receptors (OX1 and OX2). Selective OX1 receptor antagonists (SO1RA) are gaining interest for their potential use in the treatment of CNS disorders, including substance abuse, eating, obsessive compulsive, or anxiety disorders. While blocking OX2 reduces wakefulness, the expected advantage of selectively antagonizing OX1 is the ability to achieve clinical efficacy without the promotion of sleep. Herein we report our discovery efforts starting from a dual orexin receptor antagonist and describe a serendipitous finding that triggered a medicinal chemistry program that culminated in the identification of the potent SO1RA ACT-539313. Efficacy in a rat model of schedule-induced polydipsia supported the decision to select the compound as a preclinical candidate. Nivasorexant (20) represents the first SO1RA to enter clinical development and completed a first proof of concept phase II clinical trial in binge eating disorder in 2022.

Exploring structural determinants of neuroprotection bias on novel glypromate conjugates with bioactive amines.

Neurodegenerative disorders of the central nervous system (CNS) such as Alzheimer's and Parkinson's diseases, afflict millions globally, posing a significant public health challenge. Despite extensive research, a critical hurdle in effectively treating neurodegenerative diseases is the lack of neuroprotective drugs that can halt or reverse the underlying disease processes. In this work, we took advantage of the neuroprotective properties of the neuropeptide glycyl-l-prolyl-l-glutamic acid (Glypromate) for the development of new peptidomimetics using l-pipecolic acid as a proline surrogate and exploring their chemical conjugation with relevant active pharmaceutical ingredients (API) via a peptide bond. Together with prolyl-based Glypromate conjugates, a total of 36 conjugates were toxicologically and biologically evaluated. In this series, the results obtained showed that a constrained ring (l-proline) at the central position of the peptide motif accounts for enhanced toxicological profiles and biological effects using undifferentiated and differentiated human neuroblastoma SH-SY5Y cells. Additionally, it was shown that biased biological responses are API-dependent. Conjugation with (R)-1-aminoindane led to a 38-43% reduction of protein aggregation induced by Aß25-35 (10 µM), denoting a 3.2-3.6-fold improvement in comparison with the parent neuropeptide, with no significative difference between functionalization at α and γ-carboxyl ends. On the other hand, the best-performing neuroprotective conjugate against the toxicity elicited by 6-hydroxydopamine (6-OHDA, 125 µM) was obtained by conjugation with memantine at the α-carboxyl end, resulting in a 2.3-fold improvement of the neuroprotection capacity in comparison with Glypromate neuropeptide. Altogether, the chemical strategy explored in this work shows that the neuroprotective capacity of Glypromate can be modified and fine-tuned, opening a new avenue for the development of biased neurotherapeutics for CNS-related disorders.

Neuritin Alleviates Diabetic Retinopathy by Regulating Endoplasmic Reticulum Stress in Rats.

BACKGROUND: Neuritin, a small-molecule neurotrophic factor, maintains neuronal cell activity, inhibits apoptosis, promotes process growth, and regulates neural progenitor cell differentiation, migration, and synaptic maturation. Neuritin helps retinal ganglion cells (RGCs) survive optic nerve injury in rats and regenerate axons. However, the role of Neuritin in Diabetic retinopathy (DR) is unclear. OBJECTIVE: This study is intended to investigate the effect and mechanism of Neuritin in DR. For this purpose, we established DR rat models and injected Neuritin into them. This study provides a potential treatment for diabetic retinopathy. METHODS: The rat model of DR was established by streptozotocin (STZ) injection, and the effect of Neuritin on DR was detected by intravitreal injection. Histological analysis was performed by H&E and TUNEL methods. The mRNA and protein expressions of endoplasmic reticulum stress (ERS) pathway-related transcription factors were detected by qRT-PCR and western blot. The blood-retinal barrier (BRB) function was assessed using the patch-clamp technique and Evans blue leakage assay. RESULTS: Neuritin significantly improved the retinal structure, restrained the apoptosis of retinal cells, and protected the normal function of BRB in DR model rats. Mechanistically, Neuritin may function by inhibiting the expression of GRP78, ASK1, Caspase-12, VEGF, and so on. CONCLUSION: Our results indicate that Neuritin alleviates retinal damage in DR rats via the inactive endoplasmic reticulum pathway. Our study provides a potential treatment for DR.

Orexin deficiency modulates the dipsogenic effects of angiotensin II in a sex-dependent manner.

The orexin (hypocretin) neuropeptide system is an important regulator of ingestive behaviors, i.e., it promotes food and water intake. Here, we investigated the role of orexin in drinking induced by the potent dipsogen angiotensin II (ANG II). Specifically, male and female orexin-deficient mice received intracerebroventricular (ICV) injections of ANG II, followed by measuring their water intake within 15 min. We found that lower doses of ANG II (100 ng) significantly stimulated drinking in males but not in females, indicating a general sex-dependent effect that was not affected by orexin deficiency. However, higher doses of ANG II (500 ng) were sufficient to induce drinking in female wild-type mice, while female orexin-deficient mice still did not respond to the dipsogenic properties of ANG II. In conclusion, these results suggest sex-dependent effects in ANG II-induced drinking and further support the sexual dimorphism of orexin system functions.

The vasoactive effects of bradykinin, vasoactive intestinal peptide, calcitonin gene-related peptide and neuropeptide Y depend on the perivascular tissue in porcine retinal arterioles in vitro.

PURPOSE: The retina contains a number of vasoactive neuropeptides and corresponding receptors, but the role of these neuropeptides for tone regulation of retinal arterioles has not been studied in detail. METHODS: Porcine arterioles with preserved perivascular retinal tissue were mounted in a wire myograph, and the tone was measured after the addition of increasing concentrations of bradykinin, vasoactive intestinal peptide (VIP), neuropeptide Y (NPY), substance P (SP), calcitonin gene-related peptide (CGRP) and brain natriuretic peptide (BNP). The experiments were performed during inhibition of the synthesis of nitric oxide (NO), prostaglandins and dopamine and were repeated after removal of the perivascular retinal tissue. RESULTS: Bradykinin, VIP and CGRP induced significant concentration-dependent dilatation and NPY significant concentration-dependent contraction of the arterioles in the presence of perivascular retinal tissue (p < 0.03 for all comparisons) but not on isolated arterioles. BNP and SP had no effect on vascular tone. The NOS inhibitor L-NAME reduced bradykinin- and VIP-induced relaxation (p < 0.001 for both comparisons), whereas none of the other inhibitors influenced the vasoactive effects of the studied neuropeptides. CONCLUSION: The effects of neuropeptides on the tone of retinal arterioles depend on the perivascular retinal tissue and may involve effects other than those mediated by nitric oxide, prostaglandins and adrenergic compounds. Investigation of the mechanisms underlying the vasoactive effect of neuropeptides may be important for understanding and treating retinal diseases where disturbances in retinal flow regulation are involved in the disease pathogenesis.

Brain Neuropeptides, Neuroinflammation, and Irritable Bowel Syndrome.

BACKGROUND: Irritable bowel syndrome (IBS) is a functional bowel disorder characterized by chronic abdominal symptoms, but its pathogenesis is not fully understood. SUMMARY: We have recently shown in rats that neuropeptides such as orexin, ghrelin, and oxytocin act in the brain to improve the intestinal barrier dysfunction, which is a major pathophysiology of IBS. We have additionally shown that the neuropeptides injected intracisternally induced a visceral antinociceptive action against colonic distension. Since it has been known that intestinal barrier dysfunction causes visceral hypersensitivity, the other main pathophysiology of IBS, the neuropeptides act centrally to reduce leaky gut, followed by improvement of visceral sensation, leading to therapeutic action on IBS. It has been recently reported that there is a bidirectional relationship between neuroinflammation in the brain and the pathophysiology of IBS. For example, activation of microglia in the brain causes visceral hypersensitivity. Accumulating evidence has suggested that orexin, ghrelin, or oxytocin could improve neuroinflammation in the CNS. All these results suggest that neuropeptides such as orexin, ghrelin, and oxytocin act in the brain to improve intestinal barrier function and visceral sensation and also induce a protective action against neuroinflammation in the brain. KEY MESSAGES: We therefore speculated that orexin, ghrelin, or oxytocin in the brain possess dual actions, improvement of visceral sensation/leaky gut in the gut, and reduction of neuroinflammation in the brain, thereby inducing a therapeutic effect on IBS in a convergent manner.

Investigating postsynaptic effects of a Drosophila neuropeptide on muscle contraction.

The Drosophila neuropeptide, DPKQDFMRFamide, was previously shown to enhance excitatory junctional potentials (EJPs) and muscle contraction by both presynaptic and postsynaptic actions. Since the peptide acts on both sides of the synaptic cleft, it has been difficult to examine postsynaptic modulatory mechanisms, particularly when contractions are elicited by nerve stimulation. Here, postsynaptic actions are examined in 3rd instar larvae by applying peptide and the excitatory neurotransmitter, l-glutamate, in the bathing solution to elicit contractions after silencing motor output by removing the central nervous system (CNS). DPKQDFMRFamide enhanced glutamate-evoked contractions at low concentrations (EC50 1.3 nM), consistent with its role as a neurohormone, and the combined effect of both substances was supra-additive. Glutamate-evoked contractions were also enhanced when transmitter release was blocked in temperature-sensitive (Shibire) mutants, confirming the peptide's postsynaptic action. The peptide increased membrane depolarization in muscle when co-applied with glutamate, and its effects were blocked by nifedipine, an L-type channel blocker, indicating effects at the plasma membrane involving calcium influx. DPKQDFMRFamide also enhanced contractions induced by caffeine in the absence of extracellular calcium, suggesting increased calcium release from the sarcoplasmic reticulum (SR) or effects downstream of calcium release from the SR. The peptide's effects do not appear to involve calcium/calmodulin-dependent protein kinase II (CaMKII), previously shown to mediate presynaptic effects. The approach used here might be useful for examining postsynaptic effects of neurohormones and cotransmitters in other systems.NEW & NOTEWORTHY Distinguishing presynaptic and postsynaptic effects of neurohormones is a long-standing challenge in many model organisms. Here, postsynaptic actions of DPKQDFMRFamide are demonstrated by assessing its ability to potentiate contractions elicited by direct application of the neurotransmitter, glutamate, when axons are silent and when transmitter release is blocked. The peptide acts at multiple sites to increase contraction, increasing glutamate-induced depolarization at the cell membrane, acting on L-type channels, and acting downstream of calcium release from the sarcoplasmic reticulum.

Neuropeptide stimulation of physiological and immunological responses in precision-cut lung slices.

Organotypic lung slices, sometimes known as precision-cut lung slices (PCLS), provide an environment in which numerous cell types and interactions can be maintained outside the body (ex vivo). PCLS were maintained ex vivo for up to a week and demonstrated health via the presence of neurons, maintenance of tissue morphology, synthesis of mucopolysaccharides, and minimal cell death. Multiple phenotypes of neuronal fibers were present in lung slices with varied size, caliber, and neurotransmitter immunoreactivity. Of the neuropeptides present in fibers, calcitonin gene-related peptide (CGRP) was the most prevalent. Exposing PCLS to recombinant CGRP resulted in the proliferation and dispersion of CD19+ B cells in slices taken selectively from females. The number of granules containing immunoreactive (ir) surfactant protein C (SPC), which are representative of alveolar type 2 cells, increased in slices from females within 24 h of exposure to CGRP. Additionally, ir-SPC granule size increased in slices from males and females across 48 h of exposure to CGRP. Exposure of PCLS to exogenous CGRP did not alter the number of solitary pulmonary neuroendocrine cells (PNEC) but did result in neuroendocrine bodies that had significantly more cells. Neuronal fiber numbers were unchanged based on ir-peripherin; however, ir-CGRP became non-detectable in fibers while unchanged in PNECs. The effects of exogenous CGRP provide insight into innate immune and neuroendocrine responses in the lungs that may be partially regulated by neural fibers. The sex-dependent nature of these changes may point to the basis for sex-selective outcomes among respiratory diseases.

RTI-263, a biased neuropeptide S receptor agonist that retains an anxiolytic effect, attenuates cocaine-seeking behavior in rats.

Neuropeptide S (NPS) is a neuromodulatory peptide that acts via a G protein-coupled receptor. Centrally administered NPS suppresses anxiety-like behaviors in rodents while producing a paradoxical increase in arousal. In addition, NPS increases drug-seeking behavior when administered during cue-induced reinstatement. Conversely, an NPS receptor (NPSR) antagonist, RTI-118, decreases cocaine-seeking behavior. A biased NPSR ligand, RTI-263, produces anxiolytic-like effects and has memory-enhancing effects similar to those of NPS but without the increase in arousal. In the present study, we show that RTI-263 decreased cocaine seeking by both male and female rats during cue-induced reinstatement. However, RTI-263 did not modulate the animals' behaviors during natural reward paradigms, such as palatable food intake, feeding during a fasting state, and cue-induced reinstatement of sucrose seeking. Therefore, NPSR biased agonists are a potential pharmacotherapy for substance use disorder because of the combined benefits of decreased drug seeking and the suppression of anxiety.