Chronic stress and social housing differentially affect neurogenesis in male and female rats

Stress plays an important role in the development of affective disorders. Women show a higher prevalence for these disorders than men. The course of a depression is thought to be positively influenced by social support. We have used a chronic stress model in which rats received foot-shocks daily for 3 weeks. Since rats are social animals we hypothesised that 'social support' might reduce the adverse effects of chronic stress. To test this hypothesis, male and female rats were housed individually or socially in unisex groups of four rats. The proliferation marker bromodeoxyuridine (BrdU) was injected 2 weeks before the sacrifice to investigate if stress and social housing influenced the survival of proliferating cells in the dentate gyrus (DG). To investigate changes in proliferation, another group of rats was sacrificed the day after the last BrdU injection. Stress significantly decreased BrdU labelling in individually housed males and not significantly in socially housed males. In individually housed females stress increased BrdU labelling, which was prevented by social housing. The increase found in females is most likely caused by differences in survival rate, since cell proliferation was not affected by stress or housing conditions. These results indicate that social support can affect neurogenesis in both female and male rats, however in a different way.     

Alteration in dendritic morphology of pyramidal neurons from the prefrontal cortex of rats with renovascular hypertension

We have studied, in the rat, the dendritic morphological changes of the pyramidal neurons of the medial part of the prefrontal cortex induced by the chronic effect of high blood pressure. Renovascular hypertension was induced using a silver clip on the renal artery by surgery. The morphology of the pyramidal neurons from the medial part of the prefrontal cortex was investigated in these animals. The blood pressure was measured to confirm the increase in the arterial blood pressure. After 16 weeks of increase in the arterial blood pressure, the animals were sacrificed by overdoses of sodium pentobarbital and perfused intracardially with a 0.9% saline solution. The brains were removed, processed by the Golgi-Cox stain method and analyzed by the Sholl method. The dendritic morphology clearly showed that the hypertensive animals had an increase (32%) in the dendritic length of the pyramidal cells with a decrease (50%) in the density of dendritic spines when compared with sham animals. The branch-order analysis showed that the animals with hypertension exhibit more dendritic arborization at the level of the first to fourth branch order. This result suggests that renovascular hypertension may in part affect the dendritic morphology in this limbic structure, which may implicate cognitive impairment in hypertensive patients.     

The non-steroidal anti-inflammatory drug diclofenac sodium attenuates IFN-alpha induced alterations to monoamine turnover in prefrontal cortex and hippocampus

Interferon-alpha (IFN-alpha) administration induces major depression in a significant number of patients undergoing treatment for viral illnesses and other chronic diseases. Non-steroidal anti-inflammatory drugs (NSAIDs) are known to counteract a number of IFN-alpha-induced side effects, including pro-inflammatory cytokine activation and stress hormone release. To investigate this possibility further, we sought to determine the effect of the NSAID diclofenac sodium on monoamine turnover in brain induced by acute IFN-alpha exposure. Eleven male, Wistar rats (8 weeks old) were pretreated with diclofenac (20 mg/kg, s.c.) or saline, followed by intracerebroventricular (i.c.v.) infusion of IFN-alpha (1000 IU in 5 microl) or vehicle. The prefrontal cortex, striatum, and hippocampus were isolated and samples were assayed for monoamines and major metabolites by high-pressure liquid chromatography with electrochemical detection. The data show that acute IFN-alpha increased serotonin turnover in prefrontal cortex and increased dopamine turnover in hippocampus, while pre-treatment with diclofenac completely prevented these neurochemical responses. Importantly, these changes were recorded in two brain areas known to be important in depression and antidepressant action. These data offer support for a novel role of NSAIDs in modulating IFN-alpha-induced neurochemical alterations, and raise the possibility of the use of NSAIDs for the prevention of IFN-alpha-induced depression.     

Pressor effects of electrical stimulation of medial prefrontal cortex in unanesthetized rats

The medial prefrontal cortex (MPFC) is involved in central nervous system (CNS)-mediated cardiovascular modulation. We compared the cardiovascular effects of electrical stimulation (EE) of the MPFC in unanesthetized rats to those observed after stimulation of the same area in urethane-anesthetized rats. Electrical stimulation (35, 106, 177, 247, 318, and 389 microA rms/10 sec, 60-Hz sine wave) of the MPFC of urethane-anesthetized rats caused depressor responses of stimulus-related intensity. The cardiovascular response to electrical stimulation of the MPFC in unanesthetized rats was characterized by stimulus-related pressor responses. No significant heart rate changes were observed during the EE period in any case. The pressor response to electrical stimulation (106 microA rms/10 sec, 60-Hz sine wave) of the MPFC was not affected by intravenous pretreatment with the vasopressin antagonist dTyr(CH(2))(5)(Me)AVP (50 microg/kg, intravenously), by hypophysectomy, or by intravenous pretreatment with the angiotensin II antagonist losartan (1 mg/kg, intravenously). The pressor response was blocked by intravenous pretreatment with the ganglionic blocker mecamylamine (2 mg/kg, intravenously) but was not affected by adrenal demedullation, thus suggesting involvement of the neural component of the sympathetic nervous system without a major involvement of its hormonal component. Our results confirmed the occurrence of depressor responses after electrical stimulation of the MPFC in urethane-anesthetized rats and evidenced that only pressor responses are observed after its stimulation in unanesthetized rats. The fact that the pressor response to the stimulation of the MPFC was blocked by a ganglioplegic suggests that the MPFC has functional excitatory actions over the sympathetic nervous system.     

Alterations in QT dispersion in the surface electrocardiogram of female adolescents diagnosed with restricting-type anorexia nervosa

OBJECTIVE: QT dispersion (QTd), defined as the difference between the longest interval and the shortest interval in the 12-lead electrocardiogram (ECG), is a measure of myocardial repolarization inhomogeneity. We assessed QTd in malnourished anorexia nervosa (AN) inpatients and following weight restoration. METHODS: QTd analysis, anthropometric evaluations, and laboratory tests were carried out in 30 malnourished female adolescent AN restricting-type (AN-R) inpatients and following weight restoration. RESULTS: A significant increase was found in weight/height ratio and body mass index from malnourished stage to weight restoration, paralleled by a significant decrease in QTd (70+/-16 vs. 47+/-16 ms; P<.0001). No correlations were found between ECG indices and anthropometric and laboratory measures. CONCLUSION: Elevated QTd in malnourished AN-R inpatients may indicate possible cardiac autonomic imbalance and/or myocardial damage, likely corrected following weight restoration.     

Increase in vulnerability of atrial fibrillation in an acute intermittent hypoxia model: Importance of autonomic imbalance

Background

The underlying mechanism of atrial fibrillation (AF) in association with hypoxia remains unclear. This study aimed to investigate the AF vulnerability in an acute intermittent hypoxia model and ascertained the mechanism.

Methods and results

In 15 anesthetized open-chest dogs (10 experimental and 5 control), multiple-electrode catheters were sutured to the surface of atria, pulmonary veins and superior vena cava. The ventilators were adjusted to simulate the intermittent hypoxia for 1 h. The heart rate, blood pressure, blood gas, heart rate variability (HRV), effective refractory period (ERP) and window of vulnerability (WOV) as a measure of AF inducibility were measured. As the hypoxia prolonged, the pH, PaO₂ and SaO₂ progressively decreased, while the PaCO₂ increased (P < 0.05 for all). In the first 30 min of hypoxia, the HRV indexes associated with sympathetic activity (LF) and parasympathetic activity (HF) both increased but the ratio of LF/HF stayed unchanged. The ERP and WOV were not affected. In the second 30 min of hypoxia, the parasympathetic activity (HF) further increased while the ratio of LF/HF subsequently decreased. Meanwhile, the ERP shortened, and the WOV significantly increased (P < 0.05 for all). All the changes that were caused by hypoxia were reversed by 1-h re-ventilation.

Conclusions

AF vulnerability increases in the acute intermittent hypoxia model in which autonomic imbalance may play an important role. The treatment targeting autonomic imbalance may be considered when encountering this kind of AF such as in sleep apnea.     

Autonomic modulation of insulin levels in foetal sheep

The autonomic nervous system plays an important part in metabolic and circulatory adaptation of the foetus to changes in its intrauterine environment and after delivery. Foetal and neonatal glucose metabolism and insulin secretion are influenced by changes in humoral catecholamine levels as they may occur during asphyxia. To assess the role of neuronal and humoral sympathetic activity in foetal endocrine pancreatic regulation, chronically catheterized foetal sheep near term were chemically sympathectomized with 6-hydroxydopamine. Experiments were carried out in unanaesthetized foetal sheepin utero in the absence of uterine contractions. Insulin and glucose levels, blood gases, acid-base status and catecholamines were measured before, during and after a 2 min occlusion of uterine blood flow caused by mechanical constriction of the maternal aorta. Pancreatic blood flow was determined using radioactive labelled microspheres. During normoxaemia, insulin levels, pancreatic blood flow and glucose transport to the organ in sympathectomized foetuses were elevated compared with intact animals, whereas glucose concentrations did not show any significant differences. After the onset of asphyxia humoral catecholamine levels rose significantly in both groups. Insulin concentrations in the plasma of both intact and symphathectomized foetuses were no longer different indicating both indirect (blood flow and humoral catecholamine related) and direct (neurally mediated) sympathetic effects on pancreatic beta cells.     

Expression of c-fos and BDNF mRNA in subregions of the prefrontal cortex of male and female rats after acute uncontrollable stress

Women exhibit higher lifetime prevalences of stress-related disorders than men. These disorders have been associated with changes in prefrontal cortex structure and function. Here, we examine the effects of acute inescapable stress, an animal model of behavioral depression and post-traumatic stress disorder, on plasma corticosterone (CORT) and on c-fos mRNA and brain-derived neurotrophic factor (BDNF) mRNA in regions of the prefrontal and frontal cortex in male and cycling female rats. Inescapable stress consisted of 100 1 mA tailshocks, and no-stress controls remained in their home cages. Rats were sacrificed immediately (0 min) or 60 min after termination of the stressor. CORT levels were increased at both 0 and 60 min post-stress termination relative to controls, and the increase was greater in females at both time points. c-fos mRNA expression increased at 0 min in prefrontal cortical regions, but this increase was greater in males than estrus and proestrus females. At 60 min, c-fos mRNA levels were lower than at 0 min in males but not females. No correlations between CORT and c-fos mRNA levels in prefrontal regions were observed in females in the stress groups, but significant correlations were observed in males in several prefrontal regions. BDNF mRNA expression was greater in control females than control males. Inescapable stress increased BDNF mRNA expression at 0 but not 60 min in males, but there was no effect of inescapable stress on BDNF mRNA in females. These results reveal sex differences in inescapable stress-induced gene expression that may have implications for differences in vulnerability to stress-related disorders.     

Tryptophan depletion affects the autonomic stress response in generalized social anxiety disorder

In generalized social anxiety disorder (gSAD), serotonergic dysfunctions are found, as well as abnormalities of the autonomic nervous system (ANS) in basal conditions and of the hypothalamic pituitary adrenal (HPA) axis in response to psychological challenges. These findings raise the question whether these phenomena are interrelated.Therefore we designed a study in which two groups with nine pair wise age and gender matched gSAD patients (total of 10 men and 8 women), who were successfully treated with a selective serotonin reuptake inhibitor (SSRI), underwent a tryptophan depletion challenge (TD) or a placebo condition. A TD procedure temporarily decreases serotonergic neurotransmission. In order to activate the stress system the TD/placebo challenge was combined with a public speaking task. We assessed ANS responses, as measured with the promising new marker salivary alpha-amylase (sAA), and HPA-axis responses, as measured with salivary cortisol.The most important result was that the TD group showed a significant larger sAA response to the public speaking task as compared to the placebo group, reflecting hyperresponsivity of the ANS in this group, whereas no differences were seen in cortisol responses. This suggests that in gSAD there is a vulnerability of the ANS more than the HPA-axis.     

Central urocortin activation of sympathetic-regulated energy metabolism in Wistar rats

The corticotropin-releasing hormone (CRH) system, including CRH and urocortin (UCN), is implicated in the central control of appetite and energy metabolism. Urocortin, a recently isolated neuropeptide closely related to CRH is involved in the central signaling cascade that inhibits energy intake. When administered intracerebroventricularly and intra-hypothalamically, UCN potently decreases food intake. Receptors for UCN, while widely distributed, are expressed in hypothalamic nuclei. As the hypothalamus is involved in modulating autonomic outflow, UCN may also act as a catabolic neuropeptide to facilitate energy expenditure through sympathetic-regulated thermogenesis. To test the hypothesis that UCN also enhances regulatory energy expenditure via the activation of the sympathetic nervous system, we examined whole body oxygen consumption (VO₂) and colonic temperature in male Wistar rats in response to central UCN administration. That is, the intracerebroventricular injection of 1.0 μg of UCN in male Wistar rats (n=10) significantly increased whole body oxygen consumption compared to PBS control. In addition, colonic temperature was significantly increased (Δ0.7±0.08 °C) in UCN- vs. PBS-administered rats, which was prevented by pretreatment with the ganglionic blocker chlorisondamine. These studies suggest that UCN acutely increased whole body oxygen consumption and body temperature via central activation of sympathetic outflow.     

Parasympathetic varicosity proliferation and synaptogenesis in rat eyelid smooth muscle after sympathectomy

Parasympathetic innervation to eyelid smooth muscle inhibits sympathetic neurotransmission pre-junctionally without appreciable direct post-junctional effects. However, 5 weeks after sympathectomy, parasympathetic stimulation elicits substantial cholinergically mediated contractions. This study examined ultrastructural changes accompanying the conversion to parasympathetic excitation. In intact muscles, 64±9 nerve varicosities were encountered per 10⁴ μm². Most were close to muscle cells and not fully enclosed by supporting cells. Axo–axonal synapses were observed occasionally. Two days following sympathectomy, varicosity numbers were reduced by 97% and, relative to controls, remaining varicosities were farther from muscle cells and more frequently fully enclosed by supporting cells, but contained greater numbers of small spherical and large dense vesicles. By 6 weeks post-sympathectomy, numbers of varicosities per unit muscle volume increased to 14% of controls. These varicosities differed from those at 2 days in being closer to smooth muscle cells, less frequently enclosed, and having fewer small vesicles. These findings indicate that intact eyelid smooth muscle varicosities are predominantly sympathetic, but a small number of parasympathetic varicosities are present, some of which may form pre-junctional synapses with sympathetic nerves. Between 2 days and 6 weeks post-sympathectomy, varicosities increased in number and established appositions with smooth muscle cells. This suggests that parasympathetic nerves are capable of re-innervating an atypical smooth muscle target after sympathectomy, and that parasympathetic synaptogenesis is likely to contribute to conversion from pre-junctional inhibition to post-junctional excitation after sympathectomy.     

Central representation of the sympathetic nervous system in the cerebral cortex

The sympathetic-related regions of the cerebral cortex were identified in rats after pseudorabies virus injections were made in functionally different targets: adrenal gland, stellate ganglion which regulates the heart, or celiac ganglion which innervates the gastrointestinal tract. Extensive transneuronal labeling was found in limbic system areas: (1) extended amygdaloid complex, (2) lateral septum, and (3) infralimbic, insular, and ventromedial temporal cortical regions (viz., ectorhinal CORTEX=Brodmann’s area 36, perirhinal CORTEX=area 35, lateral ENTORHINAL=area 28, and ventral temporal association CORTEX=Te3 region). Deep temporal lobe structures were prominently labeled, including the amygdalopiriform and amygdalohippocampal transition areas, ventral hippocampus and ventral subiculum. The cortical circuits mediating emotional–autonomic changes (i.e., mind–body control) are discussed.     

Stress-induced changes in human salivary alpha-amylase activity -- associations with adrenergic activity

The salivary enzyme alpha-amylase has been proposed to indicate stress-reactive bodily changes. A previous study by the authors revealed marked increases in salivary alpha-amylase following psychosocial stress, indicating a stress-dependent activation of salivary alpha-amylase. Salivary alpha-amylase has been suggested to reflect catecholaminergic reactivity. Our aim was to assess/evaluate a possible relationship between salivary alpha-amylase and adrenergic parameters, i.e. catecholamines, as well as other stress markers. Using an intra-individual repeated measures design, 30 healthy young men underwent the Trier Social Stress Test (TSST), which consists of a mental arithmetic task and free speech in front of an audience and a control condition in randomized order. Salivary alpha-amylase and salivary cortisol as well as plasma catecholamines and cardiovascular activity were repeatedly measured before, during, and after both conditions. Significant differences were found between the stress and the rest condition in salivary alpha-amylase, salivary cortisol, plasma catecholamines, and cardiovascular parameters (heart rate, LF, HF, LF/HF). However, general alpha-amylase responses (area under the curve) were not associated with general responses in catecholamines and cortisol in the stress condition (r smaller than 0.25 for all analyses). Analysis of cardiovascular parameters indicates a positive relationship between amylase and sympathetic tone (LF/HF) during stress. Salivary alpha-amylase is sensitive to psychosocial stress. Since it does not seem to be closely related to other biological stress markers such as catecholamines and cortisol, salivary alpha-amylase may be a useful additional parameter for the measurement of stress.     

Increased levels of SNAP-25 and synaptophysin in the dorsolateral prefrontal cortex in bipolar I disorder

OBJECTIVE: In order to identify whether the mechanisms associated with neurotransmitter release are involved in the pathologies of bipolar disorder and schizophrenia, levels of presynaptic [synaptosomal-associated protein-25 (SNAP-25), syntaxin, synaptophysin, vesicle-associated membrane protein, dynamin I] and structural (neuronal cell adhesion molecule and alpha-synuclein) neuronal markers were measured in Brodmann's area 9 obtained postmortem from eight subjects with bipolar I disorder (BPDI), 20 with schizophrenia and 20 controls. METHODS: Determinations of protein levels were carried out using Western blot techniques with specific antibodies. Levels of mRNA were measured using real-time polymerase chain reaction. RESULTS: In BPDI, levels of SNAP-25 (p < 0.01) and synaptophysin (p < 0.05) increased. There were no changes in schizophrenia or any other changes in BPDI. Levels of mRNA for SNAP-25 were decreased in BPDI (p < 0.05). CONCLUSION: Changes in SNAP-25 and synaptophysin in BPDI suggest that changes in specific neuronal functions could be linked to the pathology of the disorder.     

Effects of sleep deprivation on cardiac autonomic and pituitary-adrenocortical stress reactivity in rats

A demanding life style, often associated with restricted time for sleep, is a growing problem in our society and may become a major health issue in the near future. Since the physiological stress system plays a critical role in coping with a challenge, it is important to know whether this system is affected by sleep loss. Although some information is available concerning the effect of sleep loss on the basal activity of the two main limbs of the stress system, the sympathetic-adrenomedullary (SAM) and the hypothalamic-pituitary-adrenocortical (HPA) axes, little is known about the effect of sleep loss on the subsequent response to a stressor. This study investigated the effects of sleep deprivation on cardiac autonomic and HPA axis (re)activity, under baseline conditions and in response to an acute emotional stressor (15-min of restraint). Rats were subjected to 48 h of sleep deprivation by placing them in slowly rotating wheels. Electrocardiographic recordings were performed via radiotelemetry and autonomic balance was quantified via time-domain indexes of heart rate variability. HPA axis activity was examined by collecting blood samples which were analyzed for plasma ACTH and corticosterone concentrations. The results show that sleep deprivation produced a tonic increase of heart rate and HPA axis activity. When the animals in a state of sleep debt were exposed to an acute restraint stress, a blunted parasympathetic antagonism was observed following sympathetic activation, together with an increased susceptibility to cardiac arrhythmias. The HPA axis response to restraint stress was also altered, but while pituitary ACTH response was attenuated, adrenal corticosterone release was unchanged, indicating an increased adrenocortical sensitivity to ACTH. The data show that sleep deprivation not only affects the baseline activity of the stress system, but it also alters its response to a subsequent stressor.     

Symbolic analysis detects alterations of cardiac autonomic modulation in congestive heart failure rats

Congestive heart failure (CHF) is associated with neurohumoral activation. Only very few studies have examined the progression of autonomic dysfunction in CHF in humans and scanty data are available in animal models of CHF. This study was performed to assess the changes in cardiac autonomic modulation during the progression of CHF in a rat model, using an innovative analysis of heart rate variability. Progression of cardiovascular autonomic dysfunction was assessed in a rat model of CHF induced by coronary artery ligation. Spectral and symbolic analyses were performed on heart period (approximated with pulse interval, PI) and systolic arterial pressure (SAP) signals, acquired ~ 2 and ~ 4 weeks after the surgical procedure. As CHF developed, symbolic analysis revealed a decrease of rhythmical physiological sympathetic modulation, as indicated by the reduction of the percentage of stable patterns. In addition, symbolic analysis revealed that runs of short–long–short and/or long–short–long PI values and high–low–high and/or low–high–low SAP values were more and more frequent as CHF progressed. On the contrary, spectral analysis of PI and SAP series was not able to detect any impairment of autonomic regulation. Indeed, low frequency and high frequency powers derived from both PI and SAP series were not significantly changed. These data indicate that the autonomic cardiovascular modulation is altered during the progression of CHF and that symbolic analysis seems to be more suitable than spectral analysis to describe alterations of heart period dynamics and of cardiovascular regulation in this animal model of CHF.     

Amygdala input to medial prefrontal cortex (mPFC) in the rat: A light and electron microscope study

This paper describes the termination pattern and synaptic connectivity of the pathway from the basolateral nucleus of the amygdala (BLA) to the medial prefrontal cortex (mPFC; areas 25, 32, and 24b) of the rat. Discrete injections of the anterograde tracer Phaseolus vulgaris-leucoagglutinin (PHA-L) were made in the BLA and detailed light microscopical observations made of the distribution of PHA-L labelled fibres and boutons in the mPFC. Labelled fibres were distributed in two tiers: predominantly within deep layer 1/layer 2 and also in layers 5/6. Fibre plexi in layers 2 and 5 were highly varicose. Electron microscopical examination of 120 labelled boutons in area 32 (60 in layer 2 and 60 in layer 5) indicated that 116 (97%) established asymmetrical synaptic contacts with dendritic spines and 4 (3%) were in synaptic contact with small dendritic shafts. No significant differences in target structures were found between layers 2 and 5. The results indicate that BLA input to mPFC in the rat predominantly innervates spine bearing dendrites in layers 2 and 5. This suggests that the neuronal operations of these processes are influenced by direct feedforward excitation from the BLA.     

Descending projections of infralimbic cortex that mediate stimulation-evoked changes in arterial pressure

The infralimbic cortex (IL) of the rat can modify autonomic nervous system activity, but the critical pathway(s) that mediate this influence are unclear. To define the potential pathways, the first series of experiments characterizes the descending projections of IL and the neighboring cortical areas using Phaseolus vulgaris leucoagglutinin (PHA-L). IL has prominent projections to the central nucleus of the amygdala (Ce), the mediodorsal nucleus of the thalamus (MD), the lateral hypothalamic area (LHA), the periaqueductal gray (PAG), the parabrachial nucleus (Pb), and the nucleus of the solitary tract (NTS). The density and selectivity of these projections suggest that the LHA and the PAG mediate the ability of the IL to regulate cardiovascular function. The second series of experiments demonstrates that locally anesthetizing neurons in either the LHA or PAG with lidocaine attenuates the hypotensive effects produced by electrical stimulation of the IL. Similarly, microinjections of cobalt chloride (a neurotransmission blocker) into the anterior portion of the LHA also decrease the arterial pressure responses to IL stimulation, suggesting that the ability of lidocaine to reversibly block the evoked response is due to inactivation of neurons in the LHA. These data indicate that hypotension evoked by stimulation of IL is mediated, at least in part, by direct or indirect projections to the LHA and through the PAG.