Early effects of lipopolysaccharide-induced inflammation on foetal brain development in rat

Studies in humans and animal models link maternal infection and imbalanced levels of inflammatory mediators in the foetal brain to the aetiology of neuropsychiatric disorders. In a number of animal models, it was shown that exposure to viral or bacterial agents during a period that corresponds to the second trimester in human gestation triggers brain and behavioural abnormalities in the offspring. However, little is known about the early cellular and molecular events elicited by inflammation in the foetal brain shortly after maternal infection has occurred. In this study, maternal infection was mimicked by two consecutive intraperitoneal injections of 200 μg of LPS (lipopolysaccharide)/kg to timed-pregnant rats at GD15 (gestational day 15) and GD16. Increased thickness of the CP (cortical plate) and hippocampus together with abnormal distribution of immature neuronal markers and decreased expression of markers for neural progenitors were observed in the LPS-exposed foetal forebrains at GD18. Such effects were accompanied by decreased levels of reelin and the radial glial marker GLAST (glial glutamate transporter), and elevated levels of pro-inflammatory cytokines in maternal serum and foetal forebrains. Foetal inflammation elicited by maternal injections of LPS has discrete detrimental effects on brain development. The early biochemical and morphological changes described in this work begin to explain the sequelae of early events that underlie the neurobehavioural deficits reported in humans and animals exposed to prenatal insults

Expression Analysis of PAC1-R and PACAP Genes in Zebrafish Embryos

This study describes the expression of the pituitary adenylate cyclase-activating polypeptide (PACAP1 and PACAP2) and PAC1 receptor genes (PAC1a-R and PAC1b-R) in the brain of zebrafish (Danio rerio) during development. In situ hybridization of the 24- and 48-hpf embryos revealed that PACAP genes were expressed in the telencephalon, the diencephalon, the rhombencephalon, and the neurons in the dorsal part of the spinal cord. PACAP2 mRNA appears to be the most abundant form during brain development. The two PAC1-R subtypes showed a similar expression pattern: mRNAs were detected in the forebrain, the thalamus, and the rhombencephalon. However, in the tectum, only PAC1b-R gene was detected. These results suggest that, in fish, PACAP may play a role in brain development

Targeting VIP and PACAP receptor signalling: new therapeutic strategies in multiple sclerosis

MS (multiple sclerosis) is a chronic autoimmune and neurodegenerative pathology of the CNS (central nervous system) affecting approx. 2.5 million people worldwide. Current and emerging DMDs (disease-modifying drugs) predominantly target the immune system. These therapeutic agents slow progression and reduce severity at early stages of MS, but show little activity on the neurodegenerative component of the disease. As the latter determines permanent disability, there is a critical need to pursue alternative modalities. VIP (vasoactive intestinal peptide) and PACAP (pituitary adenylate cyclase-activating peptide) have potent anti-inflammatory and neuroprotective actions, and have shown significant activity in animal inflammatory disease models including the EAE (experimental autoimmune encephalomyelitis) MS model. Thus, their receptors have become candidate targets for inflammatory diseases. Here, we will discuss the immunomodulatory and neuroprotective actions of VIP and PACAP and their signalling pathways, and then extensively review the structure–activity relationship data and biophysical interaction studies of these peptides with their cognate receptors

Role of PACAP and VIP Signalling in Regulation of Chondrogenesis and Osteogenesis

Pituitary adenylate cyclase activating polypeptide (PACAP) and vasoactive intestinal peptide (VIP) are multifunctional proteins that can regulate diverse physiological processes. These are also regarded as neurotrophic and anti-inflammatory substances in the CNS, and PACAP is reported to prevent harmful effects of oxidative stress. In the last decade more and more data accumulated on the similar function of PACAP in various tissues, but its cartilage- and bone-related presence and functions have not been widely investigated yet. In this summary we plan to verify the presence and function of PACAP and VIP signalling tool kit during cartilage differentiation and bone formation. We give evidence about the protective function of PACAP in cartilage regeneration with oxidative or mechanically stress and also with the modulation of PACAP signalling in vitro in osteogenic cells. Our observations imply the therapeutic perspective that PACAP might be applicable as a natural agent exerting protecting effect during joint inflammation and/or may promote cartilage regeneration during degenerative diseases of articular cartilage

Vasoactive intestinal polypeptide mediates circadian rhythmicity and synchrony in mammalian clock neurons

The mammalian suprachiasmatic nucleus (SCN) is a master circadian pacemaker. It is not known which SCN neurons are autonomous pacemakers or how they synchronize their daily firing rhythms to coordinate circadian behavior. Vasoactive intestinal polypeptide (VIP) and the VIP receptor VPAC(2) (encoded by the gene Vipr2) may mediate rhythms in individual SCN neurons, synchrony between neurons, or both. We found that Vip(−/−) and Vipr2(−/−) mice showed two daily bouts of activity in a skeleton photoperiod and multiple circadian periods in constant darkness. Loss of VIP or VPAC(2) also abolished circadian firing rhythms in approximately half of all SCN neurons and disrupted synchrony between rhythmic neurons. Critically, daily application of a VPAC(2) agonist restored rhythmicity and synchrony to VIP(−/−) SCN neurons, but not to Vipr2(−/−) neurons. We conclude that VIP coordinates daily rhythms in the SCN and behavior by synchronizing a small population of pacemaking neurons and maintaining rhythmicity in a larger subset of neurons

PACAP and VIP Modulate LPS-Induced Microglial Activation and Trigger Distinct Phenotypic Changes in Murine BV2 Microglial Cells.

Pituitary adenylate cyclase-activating polypeptide (PACAP) and vasoactive intestinal peptide (VIP) are two structurally related immunosuppressive peptides. However, the underlying mechanisms through which these peptides regulate microglial activity are not fully understood. Using lipopolysaccharide (LPS) to induce an inflammatory challenge, we tested whether PACAP or VIP differentially affected microglial activation, morphology and cell migration. We found that both peptides attenuated LPS-induced expression of the microglial activation markers Iba1 and iNOS (### p IL-1β, IL-6, Itgam and CD68 (### p < 0.001). In contrast, treatment with PACAP or VIP exerted distinct effects on microglial morphology and migration. PACAP reversed LPS-induced soma enlargement and increased the percentage of small-sized, rounded cells (54.09% vs. 12.05% in LPS-treated cells), whereas VIP promoted a phenotypic shift towards cell subpopulations with mid-sized, spindle-shaped somata (48.41% vs. 31.36% in LPS-treated cells). Additionally, PACAP was more efficient than VIP in restoring LPS-induced impairment of cell migration and the expression of urokinase plasminogen activator (uPA) in BV2 cells compared with VIP. These results suggest that whilst both PACAP and VIP exert similar immunosuppressive effects in activated BV2 microglia, each peptide triggers distinctive shifts towards phenotypes of differing morphologies and with differing migration capacities

Expression of phosphorylated cAMP response element binding protein (p-CREB) in bladder afferent pathways in VIP-/- mice with cyclophosphamide (CYP)-induced cystitis

The expression of phosphorylated cAMP response element binding protein (pCREB) in dorsal root ganglia (DRG) with and without CYP-induced cystitis (150 mg/kg, i.p; 48 hr) was determined in VIP(−/−) and wildtype (WT) mice. p-CREB-immunoreactivity (IR) was determined in bladder (Fastblue) afferent cells. Nerve growth factor (NGF) bladder content was determined by ELISAs. Basal expression of pCREB-IR in DRG of VIP(−/−) mice was (p ≤ 0.01) greater in L1, L2, L5-S1 DRG compared to WT mice. CYP treatment in WT mice increased (p ≤ 0.05) pCREB-IR in L1, L2, L5-S1 DRG. CYP treatment in VIP(−/−) mice (p ≤ 0.01) increased (p ≤ 0.01) p-CREB-IR in L6-S1 DRG compared to WT with CYP. In WT mice, bladder afferent cells (20–38%) in DRG expressed pCREB-IR under basal conditions. With CYP, pCREB-IR increased in bladder afferent cells (60–65%; L6-S1 DRG) in WT mice. In VIP(−/−) mice, bladder afferent cells (12–58%) expressed pCREB-IR under basal conditions and CYP increased pCREB expression (78–84%) in L6-S1 DRG. Urinary bladder NGF expression in VIP(−/−) mice under basal conditions or after cystitis was significantly greater than WT. Detrusor smooth muscle thickness was significantly increased in VIP(−/−) mice. Bladder NGF expression may contribute to differences in pCREB expression

Gastrointestinal dysfunction in mice with a targeted mutation in the gene encoding vasoactive intestinal polypeptide:a model for the study of intestinal ileus and Hirschsprung's disease

In 1970, Drs. Said and Mutt isolated a novel peptide from porcine intestinal extracts with powerful vasoactive properties, and named it vasoactive intestinal peptide (VIP). Since then, the biological actions of VIP in the gut as well as its signal transduction pathways have been extensively studied. A variety of in vitro and in vivo studies have indicated that VIP, expressed in intrinsic non-adrenergic non-cholinergic (NANC) neurons, is a potent regulator of gastrointestinal (GI) motility, water absorption and ion flux, mucus secretion and immune homeostasis. These VIP actions are believed to be mediated mainly by interactions with highly expressed VPAC(1) receptors and the production of nitric oxide (NO). Furthermore, VIP has been implicated in numerous physiopathological conditions affecting the human gut, including pancreatic endocrine tumors secreting VIP (VIPomas), insulin-dependent diabetes, Hirschsprung’s disease, and inflammatory bowel syndromes such as Crohn’s disease and ulcerative colitis. To further understand the physiological roles of VIP on the GI tract, we have begun to analyze the anatomical and physiological phenotype of C57BL/6 mice lacking the VIP gene. Herein, we demonstrate that the overall intestinal morphology and light microscopic structure is significantly altered in VIP(−/−) mice. Macroscopically there is an overall increase in weight, and decrease in length of the bowel compared to wild type (WT) controls. Microscopically, the phenotype was characterized by thickening of smooth muscle layers, increased villi length, and higher abundance of goblet cells. Alcian blue staining indicated that the latter cells were deficient in mucus secretion in VIP(−/−) mice. The differences became more pronounced from the duodenum to the distal jejunum or ileum of the small bowel but, became much less apparent or absent in the colon with the exception of mucus secretion defects. Further examination of the small intestine revealed larger axonal trunks and unusual unstained patches in myenteric plexus. Physiologically, the VIP(−/−) mice showed an impairment in intestinal transit. Moreover, unlike WT C57BL/6 mice, a significant percentage of VIP(−/−) mice died in the first postnatal year with overt stenosis of the gut