A Brain Region-Dependent Alteration in the Expression of Vasopressin, Corticotropin-Releasing Factor, and Their Receptors Might Be in the Background of Kisspeptin-13-Induced Hypothalamic-Pituitary-Adrenal Axis Activation and Anxiety in Rats

Previously, we reported that intracerebroventricularly administered kisspeptin-13 (KP-13) induces anxiety-like behavior and activates the hypothalamic-pituitary-adrenal (HPA) axis in rats. In the present study, we aimed to shed light on the mediation of KP-13′s stress-evoking actions. The relative gene expressions of the corticotropin-releasing factor (Crf, Crfr1, and Crfr2) and arginine vasopressin (Avp, Avpr1a, and Avpr1b) systems were measured in the amygdala and hippocampus of male Wistar rats after icv KP-13 treatment. CRF and AVP protein content were also determined. A different set of animals received CRF or V1 receptor antagonist pretreatment before the KP-13 challenge, after which either an open-field test or plasma corticosterone levels measurement was performed. In the amygdala, KP-13 induced an upregulation of Avp and Avpr1b expression, and a downregulation of Crf. In the hippocampus, the mRNA level of Crf increased and the level of Avpr1a decreased. A significant rise in AVP protein content was also detected in the amygdala. KP-13 also evoked anxiety-like behavior in the open field test, which the V1 receptor blocker antagonized. Both CRF and V1 receptor blockers reduced the KP-13-evoked rise in the plasma corticosterone level. This suggests that KP-13 alters the AVP and CRF signaling and that might be responsible for its effect on the HPA axis and anxiety-like behavior

The interaction of Urocortin II and Urocortin III with amygdalar and hypothalamic cotricotropin-releasing factor (CRF) - reflections on the regulation of the hypothalamic-pituitary-adrenal (HPA) axis

Electroosmotic pumps employing silica frits synthesized from potassium silicate as a stationary phase show strong electroosmotic flow velocity and resistance to pressure-driven flow. We characterize these pumps and measure an electroosmotic mobility of 2.5x10(-8) m(2)/V s and hydrodynamic resistance per unit length of 70 x10(17) Pa s/m(4) with a standard deviation of less than 2% even when varying the amount of water used in the potassium silicate mixture. Furthermore, we demonstrate the simple integration of these pumps into a proofof- concept PDMS lab-on-a-chip device fabricated from a 3D-printed template.Funding Agencies|Vetenskapsradet [2007-3983, 2008-7537, 2011-6404]</p

The effects of CRF and urocortins on the sociability of mice

The aim of our study was to determine the role of corticotropin-releasing factor (CRF), the urocortins (Ucn1, Ucn2 and Ucn3) and their receptors (CRF1 and CRF2) in the sociability of mice. Male CFLP mice were administered intracerebroventricularly (icv) with CRF and urocortins alone or in combination with antalarmin (specific CRF1 antagonist) and astressin(2B) (specific CRF2 antagonist) and then investigated in a Crawley social interaction test arena, that consists of three chambers. An unknown male in a cage was put in the first chamber and an empty cage was put in the opposite chamber. The tested male was habituated with the middle chamber for 5 min and then allowed to explore the remaining chambers for 5 min, during which the number of entries and the time of interaction were measured. Intracerebroventricular administration of CRF decreased significantly the number of entries and the time of interaction with the unknown male and these effects were blocked by antalarmin, but not astressin2B. In contrast, central administration of Ucn1 increased significantly the number of entries into the chamber of the unknown male, without changing the time of interaction and this effect was blocked by astressin2B, but not antalarmin. Central administration of Ucn2 and Ucn3 didn't influence remarkably the number of entries, but it reduced the time of interaction between the male mice. Our study suggests that CRF and Ucn1 may play important, but different roles in sociability, and that Ucn2 and Ucn3, playing similar roles, must be also involved in social interactions. (C) 2017 Elsevier B.V. All rights reserved

Chronic kidney disease may evoke anxiety by altering CRH expression in the amygdala and tryptophan metabolism in rats

Chronic kidney disease (CKD) is associated with anxiety; however, its exact mechanism is not well understood. Therefore, the aim of the present study was to assess the effect of moderate CKD on anxiety in rats. 5/6 nephrectomy was performed in male Wistar rats. 7 weeks after, anxiety-like behavior was assessed by elevated plus maze (EPM), open field (OF), and marble burying (MB) tests. At weeks 8 and 9, urinalysis was performed, and blood and amygdala samples were collected, respectively. In the amygdala, the gene expression of Avp and the gene and protein expression of Crh , Crhr1 , and Crhr2 were analyzed. Furthermore, the plasma concentration of corticosterone, uremic toxins, and tryptophan metabolites was measured by UHPLC-MS/MS. Laboratory tests confirmed the development of CKD. In the CKD group, the closed arm time increased; the central time and the total number of entries decreased in the EPM. There was a reduction in rearing, central distance and time in the OF, and fewer interactions with marbles were detected during MB. CKD evoked an upregulation of gene expression of Crh , Crhr1 , and Crhr2 , but not Avp , in the amygdala. However, there was no alteration in protein expression. In the CKD group, plasma concentrations of p-cresyl-sulfate, indoxyl-sulfate, kynurenine, kynurenic acid, 3-hydroxykynurenine, anthranilic acid, xanthurenic acid, 5-hydroxyindoleacetic acid, picolinic acid, and quinolinic acid increased. However, the levels of tryptophan, tryptamine, 5-hydroxytryptophan, serotonin, and tyrosine decreased. In conclusion, moderate CKD evoked anxiety-like behavior that might be mediated by the accumulation of uremic toxins and metabolites of the kynurenine pathway, but the contribution of the amygdalar CRH system to the development of anxiety seems to be negligible at this stage

Kisspeptin modulates the activity of the stress system and associated behaviours, the body temperature and nociception

The objective of this dissertation was to explore if kisspeptins have a more wide- spread function in the CNS then the regulation of the reproductive axis. Kisspeptin, a member of the RF-amide family, is an endogenous neurohormone responsible for the organisation of the HPG axis. As the product of the Kiss-1 gene it is present in four biologically active forms consisting of 54, 14, 13 and 10 amino acids. Both the distribution data of the peptides and its cognant receptor, both the already well- established function of related RF-amide peptides in nociception and neuroendocrine processes, as well as the emerging evidence associating kisspeptins with metabolic integration underlies the need for further studies. Therefore, we have investigated if centrally injected kisspeptin has any role in the organisation of the stress response and stress-associated behaviours, general activity and thermoregulation. We have also sought to know whether kisspeptin, similarly to other RF-amide peptides can modulate pain sensitivity and can impact the acute actions of morphine on nocicep- tion. KP-13 was administered icv. in different doses to adult male Sprague-Dawley rats, the behaviour of which was then observed by means of telemetry, OF and EPM tests. Additionally, plasma concentrations of corticosterone were measured in order to assess the influence of KP-13 on the HPA system. The effects on core temperature were monitored continuously via telemetry. To assess the mediation of KP’s effect on EPM behaviour AVP1R or KISS1R antagonists were administered 30 min before peptide treatment, whereas in the corticosterone measurements AVP1R antagonist and α-helical CRF(9-41) pretreatments were applied. In another set of experiments antidepressant-like effects of KP-13 were studied and the potential involvement of the adrenergic, serotonergic, cholinergic, dopaminergic and gabaergic receptors in its antidepressant-like effects was investigated in a modified FST in mice. The mice were pretreated with a non-selective α-adrenergic receptor (AR) antagonist, phenoxybenzamine, an α1/α2β-AR antagonist, prazosin, an α2-AR antagonist, yohimbine, a α-AR antagonist, propranolol, a mixed 5 − HT1/5 − HT2 serotonergic receptor antagonist, methysergide, a nonselective 5 − HT2 serotonergic receptor antagonist, cyproheptadine, a nonselective muscarinic acetylcholine receptor antagonist, atropine, a D2, D3, D4 dopamine receptor antagonist, haloperidol, or a γ-aminobutyric acid subunit A (GABA-A) receptor antagonist, bicuculline. Finally, in the third study we have endeavoured to shed light on the possible interaction of kisspeptin with morphine on nociception in adult male mice. Following the icv. administration of KP-13, the pain sensitivity was measured by a heat-radiant tail flick test. To assess the effect of KP-13 on acute morphine analgesia, the most effective dose was injected 30 minutes before a single subcutaneous dose of morphine Page 46 (2.4 mg/kg). Furthermore, acute morphine tolerance was evaluated by giving a bolus injection of morphine (60 mg/kg) 24 h before the pain sensitivity to a challenge dose of morphine (4 mg/kg) was measured. To evaluate if KP-13 has an effect on withdrawal signs, 3 h after the development of acute morphine tolerance naloxone- precipitated withdrawal was inducted and stereotyped jumping behaviour, weight and body temperature changes were observed. Our results demonstrated that KP-13 stimulated the horizontal locomotion in the OF test and decreased the number of entries into and the time spent in the open arms during the EPM tests, which was blocked by both the AVP1R and KISS1R antagonists. The peptide also caused marked elevations in the spontaneous locomotor activity and the core temperature recorded by the telemetric system, and significantly increased the basal corticosterone level, the last of which was prevented by again the AVP1R blocker. The FST revealed that KP-13 reversed the immobility, climbing and swimming times, suggesting antidepressant-like effects. Phenoxybenzamine, yohimbine and cyproheptadine prevented the effects of KP-13 on the immobility, climbing and swim- ming times, whereas prazosin, propranolol, methysergide, atropine, haloperidol and bicuculline did not modify the effects of KP-13. Furthermore, our results showed that KP-13 significantly decreased the pain threshold. Peptide treatment also depressed the acute nociceptive effect of morphine and attenuated the development of morphine tolerance. In the withdrawal experi- ment KP-13 was found to exacerbate withdrawal signs, however our results were not significant..

Obestatin prevents analgesic tolerance to morphine and reverses the effects of mild morphine withdrawal in mice

Obestatin is a 23-amino acid gut-derived neuropeptide, encoded by the same gene with ghrelin. The goal of this study was to examine the effects of obestatin on the acute and chronic analgesic actions of morphine and on mild morphine withdrawal. Open-field (OF) and elevated plus maze (EPM) tests were used to assess mild morphine withdrawal-induced behavior changes and the heat-radiant tail-flick assay was used to investigate analgesic actions of morphine. CFLP male mice were treated twice a day with graded doses of morphine in EPM and OF experiments and once a day in tail-flick studies. Obestatin (1.5mug/2mul) was administrated once a day in all experiments. Furthermore, 0.2mg/kg naloxone or saline was administered after the final injection of morphine at a dose of 20mg/kg in EPM and OF. These behavioral parameters were monitored in the OF: the percentage of center ambulation time and distance; whereas in the EPM: the time spent in open arms and the entries into open arms compared to the total time (%OAT) and entries (%OAE). In the OF, obestatin significantly decreased the percentage of time spent in the center in mice undergoing naloxone-precipitated mild morphine withdrawal. EPM results were similar to open field, but obestatin had no significant effect on parameters mentioned above. Besides, obestatin maintained the analgesic effect of morphine 90 and 120min after morphine injection in mice treated with morphine receiving obestatin compared to mice treated with morphine. In tolerance studies, obestatin diminished the analgesic tolerance to morphine on the 5th day. In this study we confirmed that obestatin reversed the effect of mild morphine withdrawal and enhances the analgesic effect of morphine. These data suggest that obestatin may have a role in opioid-induced analgesia and in behavioral responses induced by opioid withdrawal