Evelin Painsipp

Reduced anxiety‐like and depression‐related behavior in neuropeptide Y Y4 receptor knockout mice

Neuropeptide Y (NPY) acting through Y1 receptors reduces anxiety‐ and depression‐like behavior in rodents, whereas Y2 receptor stimulation has the opposite effect. This study addressed the implication of Y4 receptors in emotional behavior by comparing female germ line Y4 knockout (Y4−/−) mice with control and germ line Y2−/− animals. Anxiety‐ and depression‐like behavior was assessed with the open field (OF), elevated plus maze (EPM), stress‐induced hyperthermia (SIH) and tail suspension tests (TST), respectively. Learning and memory were evaluated with the object recognition test (ORT). In the OF and EPM, both Y4−/− and Y2−/− mice exhibited reduced anxiety‐related behavior and enhanced locomotor activity relative to control animals. Locomotor activity in a familiar environment was unchanged in Y4−/− but reduced in Y2−/− mice. The basal rectal temperature exhibited diurnal and genotype‐related alterations. Control mice had temperature minima at noon and midnight, whereas Y4−/− and Y2−/− mice displayed only one temperature minimum at noon. The magnitude of SIH was related to time of the day and genotype in a complex manner. In the TST, the duration of immobility was significantly shorter in Y4−/− and Y2−/− mice than in controls. Object memory 6 h after initial exposure to the ORT was impaired in Y2−/− but not in Y4−/− mice, relative to control mice. These results show that genetic deletion of Y4 receptors, like that of Y2 receptors, reduces anxiety‐like and depression‐related behavior. Unlike Y2 receptor knockout, Y4 receptor knockout does not impair object memory. We propose that Y4 receptors play an important role in the regulation of behavioral homeostasis.

Deletion of the acid-sensing ion channel ASIC3 prevents gastritis-induced acid hyperresponsiveness of the stomach-brainstem axis.

&NA; Gastric acid challenge of the rat and mouse stomach is signalled to the brainstem as revealed by expression of c‐Fos. The molecular sensors relevant to the detection of gastric mucosal acidosis are not known. Since the acid‐sensing ion channels ASIC2 and ASIC3 are expressed by primary afferent neurons, we examined whether knockout of the ASIC2 or ASIC3 gene modifies afferent signalling of a gastric acid insult in the normal and inflamed stomach. The stomach of conscious mice (C57BL/6) was challenged with intragastric HCl; two hours later the activation of neurons in the nucleus tractus solitarii (NTS) of the brainstem was visualized by c‐Fos immunocytochemistry. Mild gastritis was induced by addition of iodoacetamide (0.1%) to the drinking water for 7 days. Exposure of the gastric mucosa to HCl (0.25 M) caused a 3‐fold increase in the number of c‐Fos‐positive neurons in the NTS. This afferent input to the NTS remained unchanged by ASIC3 knockout, whereas ASIC2 knockout augmented the c‐Fos response to gastric HCl challenge by 33% (P < 0.01). Pretreatment of wild‐type mice with iodoacetamide induced mild gastritis, as revealed by increased myeloperoxidase activity, and enhanced the number of NTS neurons responding to gastric HCl challenge by 41% (P < 0.01). This gastric acid hyperresponsiveness was absent in ASIC3 knockout mice but fully preserved in ASIC2 knockout mice. The current data indicate that ASIC3 plays a major role in the acid hyperresponsiveness associated with experimental gastritis. In contrast, ASIC2 appears to dampen acid‐evoked input from the stomach to the NTS.

Sex-dependent control of murine emotional-affective behaviour in health and colitis by peptide YY and neuropeptide Y

BACKGROUND AND PURPOSE Peptide YY (PYY) and neuropeptide Y (NPY) are involved in regulating gut and brain function. Because gastrointestinal inflammation is known to enhance anxiety, we explored whether experimental colitis interacts with genetic deletion (knockout) of PYY and NPY to alter emotional‐affective behaviour.

Implication of neuropeptide-Y Y2 receptors in the effects of immune stress on emotional, locomotor and social behavior of mice

Neuropeptide Y (NPY) is involved in the regulation of emotional behavior, and there is indirect evidence for a role of NPY in the cerebral responses to peripheral immune challenge. Since the NPY receptors involved in these reactions are not known, we investigated the effect of Escherichia coli lipopolysaccharide (LPS) on emotional, locomotor and social behavior, body temperature and circulating corticosterone in female Y2 (Y2-/-) and Y4 (Y4-/-) receptor knockout mice. LPS (0.1mg/kg injected IP 2.5h before testing) increased rectal temperature in control and Y4-/- mice to a larger degree than in Y2-/- animals. Both Y2-/- and Y4-/- mice exhibited reduced anxiety-related and depression-like behavior in the open field, elevated plus-maze and tail suspension test, respectively. While depression-like behavior was not changed by LPS, anxiety-related behavior was enhanced by LPS in Y2-/-, but not control and Y4-/- animals. Y2-/- mice were also particularly susceptible to the effect of LPS to attenuate locomotor behavior and social interaction with another mouse. The corticosterone response to LPS was blunted in Y2-/- mice which presented elevated levels of circulating corticosterone following vehicle treatment. These data show that Y2-/- mice are particularly sensitive to the effects of LPS-evoked immune stress to attenuate locomotion and social interaction and to increase anxiety-like behavior, while the LPS-induced rise of temperature and circulating corticosterone is suppressed by Y2 receptor knockout. Our observations attest to an important role of endogenous NPY acting via Y2 receptors in the cerebral response to peripheral immune challenge.

Gene–environment interaction influences anxiety-like behavior in ethologically based mouse models

Ethologically based animal models are widely used; however, results from different laboratories vary significantly which may partly be due to the lack of standardization. Here, we examined the effects of circadian rhythm, lighting condition and mouse strain (BALB/c and C57BL/6, known to differ in measures of avoidance and risk assessment behavior) on two well established behavioral tests in mice: the Elevated Plus Maze (EPM) and the Open Field (OF). Parameters from both paradigms are commonly used as indices of anxiety-like behavior. BALB/c mice and C57BL/6 mice were independently tested in the morning and at night, in regular laboratory lighting and in the dark. We developed a novel method based on infrared lighting from below, coupled to respective video-tracking equipment, which facilitates standard testing of behavior interference-free in complete darkness. The two mouse strains differed in anxiety-related variables for the EPM in the dark, and for the OF in regular laboratory lighting. Moreover, BALB/c displayed greater anxiety-like behavior than C57BL/6 in the OF but less anxiety-like behavior than C57BL/6 in the EPM. Lighting condition has a major influence on both behavioral tests and this to a considerably larger extent than circadian rhythm. In addition, the lighting condition interacts strongly with the genetic background, producing discriminative differences in the anxiety-related variables depending on mouse strain and lighting condition. These results challenge the comparability of not sufficiently standardized tests of anxiety-like behavior and emphasize the need for controlling environmental variables in behavioral phenotyping.

Prolonged Depression-Like Behavior Caused by Immune Challenge: Influence of Mouse Strain and Social Environment

Immune challenge by bacterial lipopolysaccharide (LPS) causes short-term behavioral changes indicative of depression. The present study sought to explore whether LPS is able to induce long-term changes in depression-related behavior and whether such an effect depends on mouse strain and social context. LPS (0.83 mg/kg) or vehicle was administered intraperitoneally to female CD1 and C57BL/6 mice that were housed singly or in groups of 4. Depression-like behavior was assessed with the forced swim test (FST) 1 and 28 days post-treatment. Group-housed CD1 mice exhibited depression-like behavior 1 day post-LPS, an effect that leveled off during the subsequent 28 days, while the behavior of singly housed CD1 mice was little affected. In contrast, singly housed C57BL/6 mice responded to LPS with an increase in depression-like behavior that was maintained for 4 weeks post-treatment and confirmed by the sucrose preference test. Group-housed C57BL/6 mice likewise displayed an increased depression-like behavior 4 weeks post-treatment. The behavioral changes induced by LPS in C57BL/6 mice were associated with a particularly pronounced rise of interleukin-6 in blood plasma within 1 day post-treatment and with changes in the dynamics of the corticosterone response to the FST. The current data demonstrate that immune challenge with LPS is able to induce prolonged depression-like behavior, an effect that depends on genetic background (strain). The discovery of an experimental model of long-term depression-like behavior after acute immune challenge is of relevance to the analysis of the epigenetic and pathophysiologic mechanisms of immune system-related affective disorders.

Evidence from knockout mice for distinct implications of neuropeptide-Y Y2 and Y4 receptors in the circadian control of locomotion, exploration, water and food intake.

Members of the neuropeptide-Y (NPY) family acting via Y2 and/or Y4 receptors have been proposed to participate in the control of ingestive behaviour and energy homeostasis. Since these processes vary between day and night, we explored the circadian patterns of locomotor, exploratory and ingestive behaviour in mice with disrupted genes for Y2 (Y2-/-) or Y4 (Y4-/-) receptors. To this end, the LabMaster system was used and its utility for the analysis of changes in circadian activity and ingestion caused by gene knockout evaluated. Female animals, aged 27weeks on average, were housed singly in cages fitted with sensors for water and food intake and two infrared frames for recording ambulation and rearing under a 12h light/dark cycle for 4days. Relative to WT animals, diurnal locomotion, exploration, drinking and feeding were reduced, whereas nocturnal locomotion was enhanced in Y2-/- mice. In contrast, Y4-/- mice moved more but ate and drank less during the photophase, while they ate more and explored less during the scotophase. Both Y2-/- and Y4-/- mice weighed more than WT mice. These findings attest to a differential role of Y2 and Y4 receptor signalling in the circadian control of behaviours that balance energy intake and energy expenditure. These phenotypic traits can be sensitively and continuously recorded by the LabMaster system.

Decreased anxiety in mice lacking the organic cation transporter 3.

The organic cation transporter 3 (OCT3; synonymous: extraneuronal monoamine transporter, EMT, Slc22a3) encodes an isoform of the organic cation transporters and is expressed widely across the whole brain. OCTs are a family of high-capacity, bidirectional, multispecific transporters of organic cations. These also include serotonin, dopamine and norepinephrine making OCTs attractive candidates for a variety of neuropsychiatric disorders including anxiety disorders. OCT3 has been implicated in termination of monoaminergic signalling in the central nervous system. Interestingly, OCT3 mRNA is however also significantly up-regulated in the hippocampus of serotonin transporter knockout mice where it might serve as an alternative reuptake mechanism for serotonin. The examination of the behavioural phenotype of OCT3 knockout mice thus is paramount to assess the role of OCT3. We have therefore subjected mice lacking the OCT3 gene to a comprehensive behavioural test battery. While cognitive functioning in the Morris water maze test and aggression levels measured with the resident–intruder paradigm were in the same range as the respective control animals, OCT3 knockout animals showed a tendency of increased activity and were significantly less anxious in the elevated plus-maze test and the open field test as compared to their respective wild-type controls arguing for a role of OCT3 in the regulation of fear and anxiety, probably by modulating the serotonergic tone in limbic circuitries.

Experimental gastritis in mice enhances anxiety in a gender-related manner

There is a gender-related comorbidity of pain-related and inflammatory bowel diseases with psychiatric diseases. Since the impact of experimental gastrointestinal inflammation on the emotional-affective behavior is little known, we examined whether experimental gastritis modifies anxiety, stress coping and circulating corticosterone in male and female Him:OF1 mice. Gastritis was induced by adding iodoacetamide (0.1%) to the drinking water for at least 7 days. Inflammation was assessed by gastric histology and myeloperoxidase activity, circulating corticosterone determined by enzyme immunoassay, anxiety-related behavior evaluated with the elevated plus maze and stress-induced hyperthermia tests, and depression-like behavior estimated with the tail suspension test. Iodoacetamide-induced gastritis was associated with gastric mucosal surface damage and an increase in gastric myeloperoxidase activity, this increase being significantly larger in female mice than in male mice. The rectal temperature of male mice treated with iodoacetamide was enhanced, whereas that of female mice was diminished. The circulating levels of corticosterone were reduced by 65% in female mice treated with iodoacetamide but did not significantly change in male mice. On the behavioral level, iodoacetamide treatment caused a decrease in nocturnal home-cage activity, drinking and feeding. While depression-related behavior remained unaltered following induction of gastritis, behavioral indices of anxiety were significantly enhanced in female but not male mice. There was no correlation between the estrous cycle and anxiety as well as circulating corticosterone. Radiotracer experiments revealed that iodoacetamide did not readily enter the brain, the blood-brain ratio being 20:1. Collectively, these data show that iodoacetamide treatment causes gastritis in a gender-related manner, its severity being significantly greater in female than in male mice. The induction of gastritis in female mice is associated with a reduction of circulating corticosterone and an enforcement of behavioral indices of anxiety. Gastric inflammation thus has a distinct gender-dependent influence on emotional-affective behavior and its neuroendocrine control.

Synergistic effects of NOD1 or NOD2 and TLR4 activation on mouse sickness behavior in relation to immune and brain activity markers

Toll-like receptors (TLRs) and nuclear-binding domain (NOD)-like receptors (NLRs) are sensors of bacterial cell wall components to trigger an immune response. The TLR4 agonist lipopolysaccharide (LPS) is a strong immune activator leading to sickness and depressed mood. NOD agonists are less active but can prime immune cells to augment LPS-induced cytokine production. Since the impact of NOD and TLR co-activation in vivo has been little studied, the effects of the NOD1 agonist FK565 and the NOD2 agonist muramyl dipeptide (MDP), alone and in combination with LPS, on immune activation, brain function and sickness behavior were investigated in male C57BL/6N mice. Intraperitoneal injection of FK565 (0.001 or 0.003 mg/kg) or MDP (1 or 3 mg/kg) 4 h before LPS (0.1 or 0.83 mg/kg) significantly aggravated and prolonged the LPS-evoked sickness behavior as deduced from a decrease in locomotion, exploration, food intake and temperature. When given alone, FK565 and MDP had only minor effects. The exacerbation of sickness behavior induced by FK565 or MDP in combination with LPS was paralleled by enhanced plasma protein and cerebral mRNA levels of proinflammatory cytokines (IFN-γ, IL-1β, IL-6, TNF-α) as well as enhanced plasma levels of kynurenine. Immunohistochemical visualization of c-Fos in the brain revealed that NOD2 synergism with TLR4 resulted in increased activation of cerebral nuclei relevant to sickness. These data show that NOD1 or NOD2 synergizes with TLR4 in exacerbating the immune, sickness and brain responses to peripheral immune stimulation. Our findings demonstrate that the known interactions of NLRs and TLRs at the immune cell level extend to interactions affecting brain function and behavior.