Brain state-dependent responses of midbrain dopaminergic neurons to footshock under urethane anaesthesia.

For a long time, it has been assumed that dopaminergic (DA) neurons in both the ventral tegmental area (VTA) and the substantia nigra pars compacta (SNc) uniformly respond to rewarding and aversive stimuli by either increasing or decreasing their activity, respectively. This response was believed to signal information about the perceived stimuli's values. The identification of VTA&SNc DA neurons that are excited by both rewarding and aversive stimuli has led to the categorisation of VTA&SNc DA neurons into two subpopulations: one signalling the value and the other signalling the salience of the stimuli. It has been shown that the general state of the brain can modulate the electrical activity of VTA&SNc DA neurons, but it remains unknown whether this factor may also influence responses to aversive stimuli, such as a footshock (FS). To address this question, we have recorded the responses of VTA&SNc DA neurons to FSs across cortical activation and slow wave activity brain states in urethane-anaesthetised rats. Adding to the knowledge of aversion signalling by midbrain DA neurons, we report that significant proportion of VTA&SNc DA neurons can change their responses to an aversive stimulus in a brain state-dependent manner. The majority of these neurons decreased their activity in response to FS during cortical activation but switched to increasing it during slow wave activity. It can be hypothesised that this subpopulation of DA neurons may be involved in the 'dual signalling' of both the value and the salience of the stimuli, depending on the general state of the brain.

Short-Term Preexposure to Novel Enriched Environment Augments Hippocampal Ripples in Urethane-Anesthetized Mice.

Learning and memory are affected by novel enriched environment, a condition where animals play and interact with a variety of toys and conspecifics. Exposure of animals to the novel enriched environments improves memory by altering neural plasticity during natural sleep, a process called memory consolidation. The hippocampus, a pivotal brain region for learning and memory, generates high-frequency oscillations called ripples during sleep, which is required for memory consolidation. Naturally occurring sleep shares characteristics in common with general anesthesia in terms of extracellular oscillations, guaranteeing anesthetized animals suitable to examine neural activity in a sleep-like state. However, it is poorly understood whether the preexposure of animals to the novel enriched environment modulates neural activity in the hippocampus under subsequent anesthesia. To ask this question, we allowed mice to freely explore the novel enriched environment or their standard environment, anesthetized them, and recorded local field potentials in the hippocampal CA1 area. We then compared the characteristics of hippocampal ripples between the two groups and found that the amplitude of ripples and the number of successive ripples were larger in the novel enriched environment group than in the standard environment group, suggesting that the afferent synaptic input from the CA3 area to the CA1 area was higher when the animals underwent the novel enriched environment. These results underscore the importance of prior experience that surpasses subsequent physical states from the neurophysiological point of view.

CA3-CA1 long-term potentiation occurs regardless of respiration and cardiac cycle phases in urethane-anesthetized rats.

Breathing and heartbeat synchronize to each other and to brain function and affect cognition in humans. However, it is not clear how cardiorespiratory rhythms modulate such basic processes as synaptic plasticity thought to underlie learning. Thus, we studied if respiration and cardiac cycle phases at burst stimulation onset affect hippocampal long-term potentiation (LTP) in the CA3-CA1 synapse in urethane-anesthetized adult male Sprague-Dawley rats. In a between-subjects design, we timed burst stimulation of the ventral hippocampal commissure (vHC) to systole or diastole either during expiration or inspiration and recorded responses throughout the hippocampus with a linear probe. As classical conditioning in humans seems to be most efficient at expiration-diastole, we also expected LTP to be most efficient if burst stimulation was targeted to expiration-diastole. However, LTP was induced equally in all four groups and respiration and cardiac cycle phase did not modulate CA1 responses to vHC stimulation overall. This could be perhaps because we bypassed all natural routes of external influences on the CA1 by directly stimulating the vHC. In the future, the effect of cardiorespiratory rhythms on synaptic plasticity could also be studied in awake state and in other parts of the hippocampal tri-synaptic loop.

A brain cytokine-independent switch in cortical activity marks the onset of sickness behavior triggered by acute peripheral inflammation.

Systemic inflammation triggers protective as well as pro-inflammatory responses in the brain based on neuronal and/or cytokine signaling, and it associates with acutely and protractedly disrupted cognition. However, the multiple mechanisms underlying the peripheral-central inflammatory signaling are still not fully characterized. We used intraperitoneal (i.p.) injection of lipopolysaccharide (LPS) in freely moving mice with chronically implanted electrodes for recording of local field potentials (LFP) and electrocorticography (ECoG) in the hippocampus and neocortex, respectively. We show here that a sudden switch in the mode of network activity occurred in both areas starting at 10-15 min after the LPS injection, simultaneously with a robust change from exploration to sickness behavior. This switch in cortical mode commenced before any elevations in pro-inflammatory cytokines IL-1ß, TNFα, CCL2 or IL-6 were detected in brain tissue. Thereafter, this mode dominated cortical activity for the recording period of 3 h, except for a partial and transient recovery around 40 min post-LPS. These effects were closely paralleled by changes in ECoG spectral entropy. Continuous recordings for up to 72 h showed a protracted attenuation in hippocampal activity, while neocortical activity recovered after 48 h. The acute sickness behavior recovered by 72 h post-LPS. Notably, urethane (1.3 mg/kg) administered prior to LPS blocked the early effect of LPS on cortical activity. However, experiments under urethane anesthesia which were started 24 h post-LPS (with neuroinflammation fully developed before application of urethane) showed that both theta-supratheta and fast gamma CA1 activity were reduced, DG delta activity was increased, and sharp-wave ripples were abolished. Finally, we observed that experimental compensation of inflammation-induced hypothermia 24-48 h post-LPS promoted seizures and status epilepticus; and that LPS decreased the threshold of kainate-provoked seizures beyond the duration of acute sickness behavior indicating post-acute inflammatory hyperexcitability. Taken together, the strikingly fast development and initial independence of brain cytokines of the LPS-induced cortical mode, its spectral characteristics and simultaneity in hippocampus and neocortex, as well as inhibition by pre-applied urethane, strongly suggest that the underlying mechanisms are based on activation of the afferent vagus nerve and its mainly cholinergic ascending projections to higher brain areas.

Effects of isoflurane and urethane anesthetics on glutamate neurotransmission in rat brain using in vivo amperometry.

BACKGROUND: Aspects of glutamate neurotransmission implicated in normal and pathological conditions are predominantly evaluated using in vivo recording paradigms in rats anesthetized with isoflurane or urethane. Urethane and isoflurane anesthesia influence glutamate neurotransmission through different mechanisms; however, real-time outcome measures of potassium chloride (KCl)-evoked glutamate overflow and glutamate clearance kinetics have not been compared within and between regions of the brain. In order to maintain rigor and reproducibility within the literature between the two most common methods of anesthetized in vivo recording of glutamate, we compared glutamate signaling as a function of anesthesia and brain region in the rat strain most used in neuroscience. METHODS: In the following experiments, in vivo amperometric recordings of KCl-evoked glutamate overflow and glutamate clearance kinetics (uptake rate and T80) in the cortex, hippocampus, and thalamus were performed using glutamate-selective microelectrode arrays (MEAs) in young adult male, Sprague-Dawley rats anesthetized with either isoflurane or urethane. RESULTS: Potassium chloride (KCl)-evoked glutamate overflow was similar under urethane and isoflurane anesthesia in all brain regions studied. Analysis of glutamate clearance determined that the uptake rate was significantly faster (53.2%, p < 0.05) within the thalamus under urethane compared to isoflurane, but no differences were measured in the cortex or hippocampus. Under urethane, glutamate clearance parameters were region-dependent, with significantly faster glutamate clearance in the thalamus compared to the cortex but not the hippocampus (p < 0.05). No region-dependent differences were measured for glutamate overflow using isoflurane. CONCLUSIONS: These data support that amperometric recordings of KCl-evoked glutamate under isoflurane and urethane anesthesia result in similar and comparable data. However, certain parameters of glutamate clearance can vary based on choice of anesthesia and brain region. In these circumstances, special considerations are needed when comparing previous literature and planning future experiments.

The effect of urethane and MS-222 anesthesia on the electric organ discharge of the weakly electric fish Apteronotus leptorhynchus.

Urethane and MS-222 are agents widely employed for general anesthesia, yet, besides inducing a state of unconsciousness, little is known about their neurophysiological effects. To investigate these effects, we developed an in vivo assay using the electric organ discharge (EOD) of the weakly electric fish Apteronotus leptorhynchus as a proxy for the neural output of the pacemaker nucleus. The oscillatory neural activity of this brainstem nucleus drives the fish's EOD in a one-to-one fashion. Anesthesia induced by urethane or MS-222 resulted in pronounced decreases of the EOD frequency, which lasted for up to 3 h. In addition, each of the two agents caused a manifold increase in the generation of transient modulations of the EOD known as chirps. The reduction in EOD frequency can be explained by the modulatory effect of urethane on neurotransmission, and by the blocking of voltage-gated sodium channels by MS-222, both within the circuitry controlling the neural oscillations of the pacemaker nucleus. The present study demonstrates a marked effect of urethane and MS-222 on neural activity within the central nervous system and on the associated animal's behavior. This calls for caution when conducting neurophysiological experiments under general anesthesia and interpreting their results.

Comparison of the effects of two anesthetics, isoflurane and urethane, on bladder function in rats.

We compared the effects of two anesthetics, isoflurane and urethane on bladder function in rats. Arterial pressure, cystometry (CMG), and rhythmic bladder contractions (RBCs) under isovolumetric conditions, mechanosensitive single-unit afferent activities (SAAs), bladder compliance and bladder myogenic microcontractions (bladder microcontractions), and bladder blood flow, and blood and urine biochemical tests were investigated in isoflurane- or urethane-anesthetized female rats. In results of the CMG, 3/8 rats in the isoflurane group and 7/7 rats in the urethane group showed constant bladder neurogenic contractions for micturition, whereas 5/8 rats in the isoflurane group showed unstable contractions or overflow incontinence. The RBCs appeared in the urethane group but not in the isoflurane group, and SAAs in both the Aδ- and C-fibers, bladder compliance, and bladder microcontractions in the isoflurane group were higher than those in the urethane group during bladder distension. The blood biochemical test showed that the serum calcium level was higher in the isoflurane group. The mean arterial pressure and bladder blood flow were not different between the groups. The results showed that urethane anesthesia more retains bladder neurogenic contractions for micturition compared to isoflurane. In contrast, isoflurane anesthesia more retains bladder function during the storage phase compared to urethane.

Descending pathways from the superior colliculus mediating autonomic and respiratory effects associated with orienting behaviour.

The ability to discriminate competing external stimuli and initiate contextually appropriate behaviours is a key brain function. Neurons in the deep superior colliculus (dSC) integrate multisensory inputs and activate descending projections to premotor pathways responsible for orienting, attention and defence, behaviours which involve adjustments to respiratory and cardiovascular parameters. However, the neural pathways that subserve the physiological components of orienting are poorly understood. We report that orienting responses to optogenetic dSC stimulation are accompanied by short-latency autonomic, respiratory and electroencephalographic effects in awake rats, closely mimicking those evoked by naturalistic alerting stimuli. Physiological responses were not accompanied by detectable aversion or fear, and persisted under urethane anaesthesia, indicating independence from emotional stress. Anterograde and trans-synaptic viral tracing identified a monosynaptic pathway that links the dSC to spinally projecting neurons in the medullary gigantocellular reticular nucleus (GiA), a key hub for the coordination of orienting and locomotor behaviours. In urethane-anaesthetized animals, sympathoexcitatory and cardiovascular, but not respiratory, responses to dSC stimulation were replicated by optogenetic stimulation of the dSC-GiA terminals, suggesting a likely role for this pathway in mediating the autonomic components of dSC-mediated responses. Similarly, extracellular recordings from putative GiA sympathetic premotor neurons confirmed short-latency excitatory inputs from the dSC. This pathway represents a likely substrate for autonomic components of orienting responses that are mediated by dSC neurons and suggests a mechanism through which physiological and motor components of orienting behaviours may be integrated without the involvement of higher centres that mediate affective components of defensive responses. KEY POINTS: Neurons in the deep superior colliculus (dSC) integrate multimodal sensory signals to elicit context-dependent innate behaviours that are accompanied by stereotypical cardiovascular and respiratory activities. The pathways responsible for mediating the physiological components of colliculus-mediated orienting behaviours are unknown. We show that optogenetic dSC stimulation evokes transient orienting, respiratory and autonomic effects in awake rats which persist under urethane anaesthesia. Anterograde tracing from the dSC identified projections to spinally projecting neurons in the medullary gigantocellular reticular nucleus (GiA). Stimulation of this pathway recapitulated autonomic effects evoked by stimulation of dSC neurons. Electrophysiological recordings from putative GiA sympathetic premotor neurons confirmed short latency excitatory input from dSC neurons. This disynaptic dSC-GiA-spinal sympathoexcitatory pathway may underlie autonomic adjustments to salient environmental cues independent of input from higher centres.

Novel therapeutic modalities target cell signaling of Renin-Angiotensin system/NF-κB-induced cell cycle arrest and apoptosis in urethane-induced lung cancer in mice: An in vivo study.

BACKGROUND: Lung cancer has risen to the top of the list of cancer-related deaths worldwide. Aliskiren is a direct renin inhibitor. AIM: This study aims to investigate the impact of cell signaling of Renin-Angiotensin system (RAS)/NF-κB on lung cancer by investigating the potential therapeutic effects of aliskiren for lung cancer treatment in urethane-induced lung cancer in mice. METHODS: Male BALB/c mice were randomly assigned to one of five treatment groups for 150 days, including (1) normal control; (2) aliskiren (25 mg/kg/i.p) daily, (3) urethane at a dose of 1.5 g/kg (i.p) at Day 1 and 60 (nonsmall cell lung cancer[NSCLC] group) (4) NSCLC mice received carboplatin (15 mg/kg/i.p) every other day for the last 4 successive weeks and (5) NSCLC mice treated with aliskiren daily. Tumor size was determined based on blood sampling, and lungs were isolated for biochemical analysis, western blot analysis assay, and histopathological examination. RESULTS: Urethane demonstrated significant changes in all biochemical and molecular parameters and histological patterns. Aliskiren-treated mice had significantly lower levels of NF-κB p65, Bcl-2, cyclin D1, ICAM-1, MMP-2, and Nrf2, with an increase in the catalytic activity of caspase-3 due to its RAS inhibitory mechanism. The combined urethane administration with aliskiren demonstrated a significant improvement in the histopathological examination. CONCLUSION: RAS/NF-B cell signaling is a potential therapeutic target for preventing and treating lung adenocarcinoma, evidenced by the fundamental cytotoxic mechanism and attenuation of metastasis and angiogenesis induced by the treatment of NSCLC mice with aliskiren.

Impact of anesthesia and sex on sympathetic efferent and hemodynamic responses to renal chemo- and mechanosensitive stimuli.

Activation of renal sensory nerves by chemo- and mechanosensitive stimuli produces changes in efferent sympathetic nerve activity (SNA) and arterial blood pressure (ABP). Anesthesia and sex influence autonomic function and cardiovascular hemodynamics, but it is unclear to what extent anesthesia and sex impact SNA and ABP responses to renal sensory stimuli. We measured renal, splanchnic, and lumbar SNA and ABP in male and female Sprague-Dawley rats during contralateral renal infusion of capsaicin and bradykinin or during elevation in renal pelvic pressure. Responses were evaluated with a decerebrate preparation or Inactin, urethane, or isoflurane anesthesia. Intrarenal arterial infusion of capsaicin (0.1-30.0 µM) increased renal SNA, splanchnic SNA, or ABP but decreased lumbar SNA in the Inactin group. Intrarenal arterial infusion of bradykinin (0.1-30.0 µM) increased renal SNA, splanchnic SNA, and ABP but decreased lumbar SNA in the Inactin group. Elevated renal pelvic pressure (0-20 mmHg, 30 s) significantly increased renal SNA and splanchnic SNA but not lumbar SNA in the Inactin group. In marked contrast, SNA and ABP responses to every renal stimulus were severely blunted in the urethane and decerebrate groups and absent in the isoflurane group. In the Inactin group, the magnitude of SNA responses to chemo- and mechanosensory stimuli were not different between male and female rats. Thus, chemo- and mechanosensitive stimuli produce differential changes in renal, splanchnic, and lumbar SNA. Experimentally, future investigations should consider Inactin anesthesia to examine sympathetic and hemodynamic responses to renal sensory stimuli.NEW & NOTEWORTHY The findings highlight the impact of anesthesia, and to a lesser extent sex, on sympathetic efferent and hemodynamic responses to chemosensory and mechanosensory renal stimuli. Sympathetic nerve activity (SNA) and arterial blood pressure (ABP) responses were present in Inactin-anesthetized rats but largely absent in decerebrate, isoflurane, or urethane preparations. Renal chemosensory stimuli differentially changed SNA: renal and splanchnic SNA increased, but lumbar SNA decreased. Future investigations should consider Inactin anesthesia to study SNA and hemodynamic responses to renal sensory nerve activation.

Impact of anesthesia, sex, and circadian cycle on renal afferent nerve sensitivity.

Elevated renal afferent nerve (ARNA) activity or dysfunctional reno-renal reflexes via altered ARNA sensitivity contribute to hypertension and chronic kidney disease. These nerves contain mechano- and chemosensitive fibers that respond to ischemia, changes in intrarenal pressures, and chemokines. Most studies have utilized various anesthetized preparations and exclusively male animals to characterize ARNA responses. Therefore, this study assessed the impact of anesthesia, sex, and circadian period on ARNA responses and sensitivity. Multifiber ARNA recordings were performed in male and female Sprague-Dawley rats (250-400 g) and compared across decerebrate versus Inactin, isoflurane, and urethane anesthesia groups. Intrarenal artery infusion of capsaicin (0.1-50.0 µM, 0.05 mL) produced concentration-dependent increases in ARNA; however, the ARNA sensitivity was significantly greater in decerebrate versus Inactin, isoflurane, and urethane groups. Increases in renal pelvic pressure (0-30 mmHg, 30 s) produced pressure-dependent increases in ARNA; however, ARNA sensitivity was again greater in decerebrate and Inactin groups versus isoflurane and urethane. Acute renal artery occlusion (30 s) increased ARNA, but responses did not differ across groups. Analysis of ARNA responses to increased pelvic pressure between male and female rats revealed significant sex differences only in isoflurane and urethane groups. ARNA responses to intrarenal capsaicin infusion were significantly blunted at nighttime versus daytime; however, ARNA responses to increased pelvic pressure or renal artery occlusion were not different between daytime and nighttime. These results demonstrate that ARNA sensitivity is greatest in decerebrate and Inactin-anesthetized groups but was not consistently influenced by sex.NEW & NOTEWORTHY We determined the impact of anesthesia, sex, and circadian cycle on renal afferent nerve (ARNA) sensitivity to chemical and mechanical stimuli. ARNA sensitivity to renal capsaicin infusion was greatest in decerebrate > Inactin > urethane or isoflurane groups. Elevated renal pelvic pressure significantly increased ARNA; decerebrate and Inactin groups exhibited the greatest ARNA sensitivity. Sex differences in renal afferent responses were not consistently observed. Circadian cycle altered chemosensory but not mechanosensory responses.

Functional connectivity is preserved but reorganized across several anesthetic regimes.

Under anesthesia, systemic variables and CBF are modified. How does this alter the connectivity measures obtained with rs-fMRI? To tackle this question, we explored the effect of four different anesthetics on Long Evans and Wistar rats with multimodal recordings of rs-fMRI, systemic variables and CBF. After multimodal signal processing, we show that the blood-oxygen-level-dependent (BOLD) variations and functional connectivity (FC) evaluated at low frequencies (0.031-0.25 â€‹Hz) do not depend on systemic variables and are preserved across a large interval of baseline CBF values. Based on these findings, we found that most brain areas remain functionally active under any anesthetics, i.e. connected to at least one other brain area, as shown by the connectivity graphs. In addition, we quantified the influence of nodes by a measure of functional connectivity strength to show the specific areas targeted by anesthetics and compare correlation values of edges at different levels. These measures enable us to highlight the specific network alterations induced by anesthetics. Altogether, this suggests that changes in connectivity could be evaluated under anesthesia, routinely used in the control of neurological injury.

Hippocampal and lateral entorhinal cortex physiological activity during trace conditioning under urethane anesthesia.

Significant evidence shows that the acquisition of delay conditioning can occur in out-of-awareness states, such as under anesthesia. However, it is unclear to what extent and what type of conditioning animals may achieve during nonawake states. Trace conditioning is an appealing protocol to study under anesthesia, given the long empty gap separating the conditioned and unconditioned stimuli, which must be bridged for acquisition to happen. Here, we show evidence that rats develop physiological responses during the trace conditioning paradigm under anesthesia. We recorded the activity of the hippocampus (HPC) and lateral entorhinal cortex (LEC) in urethane-anesthetized rats, along with an electromyogram and an electrocardiogram. The protocol consisted of randomly presenting two distinct sound stimuli (CS- and CS+), where only one stimulus (CS+) was assigned to be trace-paired with a footshock. A trial-average analysis revealed that animals developed significant climbing heart rate activity initiating at the CS onset and persisting during the trace period. Such climbing arose for both CS- and CS+ with similar slopes but different intercepts, suggesting CS+ heart rates were typically above CS-. The power and coherence of HPC and LEC high-frequency bands (>100 Hz) significantly increased during CS presentation and trace, similarly to CS- and CS+ and insensitive to either activated or deactivated states. To the best of our knowledge, this is the first attempt to perform a trace conditioning protocol under anesthesia. Confirmation of this procedure acquisition can allow a new preparation for the exploration of brain mechanisms that bind time-discontinuous events.NEW & NOTEWORTHY Some forms of learning, such as some types of conditioning, can occur in anesthetized states. However, the extent to which memories can be formed in these states is still an open question. Here, we investigated the trace conditioning under urethane anesthesia and found heart rate, hippocampus, and lateral entorhinal cortex physiological changes to stimuli presentation. This new preparation may allow for exploration of memory acquisition of time-discontinuous events in the nonawake brain.

Urethane attenuates early neuropathology of diisopropylfluorophosphate-induced status epilepticus in rats.

Seizures can be evident within minutes of exposure to an organophosphorus (OP) agent and often progress to status epilepticus (SE) resulting in a high mortality if left untreated. Effective medical countermeasures are necessary to sustain patients suffering from OP poisoning and to mitigate the ensuing brain injury. Here, the hypothesis was tested that a single subanesthetic dose of urethane prevents neuropathology measured 24 h following diisopropylfluorophosphate (DFP)-induced SE. Adult Sprague-Dawley rats were injected with DFP to induce SE. During SE rats displayed increased neuronal activity in the hippocampus and an upregulation of immediate early genes as well as pro-inflammatory mediators. In additional experiments rats were administered diazepam (10 mg/kg, ip) or urethane (0.8 g/kg, sc) 1 h after DFP-induced SE and compared to rats that experienced uninterrupted SE. Cortical electroencephalography (EEG) and power analysis strengthen the conclusion that urethane effectively terminates SE and prevents the overnight return of seizure activity. Neurodegeneration in limbic brain regions and the seizure-induced upregulation of key inflammatory mediators present 24 h after DFP-induced SE were strongly attenuated by administration of urethane. A trivial explanation for these beneficial effects, that urethane simply reactivates acetylcholinesterase, has been ruled out. These findings indicate that, by contrast to rats administered diazepam or rats that experience uninterrupted SE, the early neuropathology after SE is prevented by subanesthetic urethane, which terminates rather than interrupts, SE.

Electrical stimulation of the ventral tegmental area evokes sleep-like state transitions under urethane anaesthesia in the rat medial prefrontal cortex via dopamine D1 -like receptors.

The role of dopamine in regulating sleep-state transitions during, both natural sleep and under anaesthesia, is still unclear. Recording in vivo in the rat mPFC under urethane anaesthesia, we observed predominantly slow wave activity (SWA) of <1 Hz in the local field potential interrupted by occasional spontaneous transitions to a low-amplitude-fast (LAF) pattern of activity. During periods of SWA, transitions to LAF activity could be rapidly and consistently evoked by electrical stimulation of the ventral tegmental area (VTA). Spontaneous LAF activity, and that evoked by stimulation of the VTA, consisted of fast oscillations similar to those seen in the rapid eye movement (REM)-like sleep state. Spontaneous and VTA stimulation-evoked LAF activity occurred simultaneously along the dorsoventral extent of all mPFC subregions. Evoked LAF activity depended on VTA stimulation current and could be elicited using either regular (25-50 Hz) or burst stimulation patterns and was reproducible upon repeated stimulation. Simultaneous extracellular single-unit recordings showed that during SWA, presumed pyramidal cells fired phasically and almost exclusively on the Up state, while during both spontaneous and VTA-evoked LAF activity, they fired tonically. The transition to LAF activity evoked by VTA stimulation depended on dopamine D1 -like receptor activation as it was almost completely blocked by systemic administration of the D1 -like receptor antagonist SCH23390. Overall, our data demonstrate that activation of dopamine D1 -like receptors in the mPFC is important for regulating sleep-like state transitions.

Long Term Evaluation of Nanofibrous, Bioabsorbable Polycarbonate Urethane Grafts for Small Diameter Vessel Replacement in Rodents.

OBJECTIVE: Biodegradable materials for in situ vascular tissue engineering could meet the increasing clinical demand for sufficient synthetic small diameter vascular substitutes in aortocoronary bypass and peripheral vascular surgery. The aim of this study was to design a new degradable thermoplastic polycarbonate urethane (dPCU) with improved biocompatibility and optimal biomechanical properties. Electrospun conduits made from dPCU were evaluated in short and long term follow up and compared with expanded polytetrafluoroethylene (ePTFE) controls. METHODS: Both conduits were investigated prior to implantation to assess their biocompatibility and inflammatory potential via real time polymerase chain reaction using a macrophage culture. dPCU grafts (n = 28) and ePTFE controls (n = 28) were then implanted into the infrarenal abdominal aorta of Sprague-Dawley rats. After seven days, one, six, and 12 months, grafts were analysed by histology and immunohistochemistry (IHC) and assessed biomechanically. RESULTS: Anti-inflammatory signalling was upregulated in dPCU conduits and increased significantly over time in vitro. dPCU and ePTFE grafts offered excellent long and short term patency rates (92.9% in both groups at 12 months) in the rat model without dilatation or aneurysm formation. In comparison to ePTFE, dPCU grafts showed transmural ingrowth of vascular specific cells resulting in a structured neovessel formation around the graft. The graft material was slowly reduced, while the compliance of the neovessel increased over time. CONCLUSION: The newly designed dPCU grafts have the potential to be safely applied for in situ vascular tissue engineering applications. The degradable substitutes showed good in vivo performance and revealed desirable characteristics such as biomechanical stability, non-thrombogenicity, and minimal inflammatory response after long term implantation.

Development of a decerebrate model for investigating mechanisms mediating viscero-sympathetic reflexes in the spinalized rat.

Autonomic dysreflexia (AD) often occurs in individuals living with spinal cord injury (SCI) and is characterized by uncontrolled hypertension in response to otherwise innocuous stimuli originating below the level of the spinal lesion. Visceral stimulation is a predominant cause of AD in humans and effectively replicates the phenotype in rodent models of SCI. Direct assessment of sympathetic responses to viscerosensory stimulation in spinalized animals is challenging and requires invasive surgical procedures necessitating the use of anesthesia. However, administration of anesthesia markedly affects viscerosensory reactivity, and the effects are exacerbated following spinal cord injury (SCI). Therefore, the major goal of the present study was to develop a decerebrate rodent preparation to facilitate quantification of sympathetic responses to visceral stimulation in the spinalized rat. Such a preparation enables the confounding effect of anesthesia to be eliminated. Sprague-Dawley rats were subjected to SCI at the fourth thoracic segment. Four weeks later, renal sympathetic nerve activity (RSNA) responses to visceral stimuli were quantified in urethane/chloralose-anesthetized and decerebrate preparations. Visceral stimulation was elicited via colorectal distension (CRD) for 1 min. In the decerebrate preparation, CRD produced dose-dependent increases in mean arterial pressure (MAP) and RSNA and dose-dependent decreases in heart rate (HR). These responses were significantly greater in magnitude among decerebrate animals when compared with urethane/chloralose-anesthetized controls and were markedly attenuated by the administration of urethane/chloralose anesthesia after decerebration. We conclude that the decerebrate preparation enables high-fidelity quantification of neuronal reactivity to visceral stimulation in spinalized rats. NEW & NOTEWORTHY In animal models commonly used to study spinal cord injury, quantification of sympathetic responses is particularly challenging due to the increased susceptibility of spinal reflex circuits to the anesthetic agents generally required for experimentation. This constitutes a major limitation to understanding the mechanisms mediating regionally specific neuronal responses to visceral activation in chronically spinalized animals. In the present study, we describe a spinalized, decerebrate rodent preparation that facilitates quantification of sympathetic reactivity in response to visceral stimuli following spinal cord injury. This preparation enables reliable and reproducible quantification of viscero-sympathetic reflex responses resembling those elicited in conscious animals and may provide added utility for preclinical evaluation of neuropharmacological agents for the management of autonomic dysreflexia.

In Vivo Study on Mechanism Underlying Increased Pharmacological Effects of Phenobarbital in Rats with Glycerol-Induced Acute Renal Failure.

The mechanism underlying the increased pharmacological effects of phenobarbital in rats with glycerol-induced acute renal failure (ARF) was examined. In the experiments, a surgical cannula was inserted in the lateral ventricle of the rats for phenobarbital infusion, and the ARF induction was performed by intramuscular administration of 50% glycerol. The onset time of anesthesia by phenobarbital was determined with the tail flick method. In addition, cerebral microsomes were prepared from excised cerebral cortices of sham and ARF rats, and the cerebral expression of the γ-aminobutyric acid (GABA)A receptor and two cation-chloride transporters, KCC2 and NKCC1, was evaluated by Western blotting, as their functions are involved in the anesthetic effects of phenobarbital. When phenobarbital was infused in the ventricle, anesthesia was induced 2.2-times faster in ARF rats than in sham rats, and there was no detectable increase in the cerebral expression of the GABAA receptor in ARF rats. It was additionally noted that the cerebral expression of KCC2 decreased, whereas that of NKCC1 was unaltered in ARF rats. These findings indicated that the anesthetic effects of phenobarbital are potentiated in ARF rats, probably due to imbalanced cerebral expression of KCC2 and NKCC1, suggesting that altered cation-chloride handling in nerve cells is associated.

Functional connectivity under six anesthesia protocols and the awake condition in rat brain.

Resting-state functional magnetic resonance imaging (rsfMRI) is a translational imaging method with great potential in several neurobiologic applications. Most preclinical rsfMRI studies are performed in anesthetized animals, but the confounding effects of anesthesia on the measured functional connectivity (FC) are poorly understood. Therefore, we measured FC under six commonly used anesthesia protocols and compared the findings with data obtained from awake rats. The results demonstrated that each anesthesia protocol uniquely modulated FC. Connectivity patterns obtained under propofol and urethane anesthesia were most similar to that observed in awake rats. FC patterns in the α-chloralose and isoflurane-medetomidine combination groups had moderate to good correspondence with that in the awake group. The FC patterns in the isoflurane and medetomidine groups differed most from that in the awake rats. These results can be directly exploited in rsfMRI study designs to improve the data quality, comparability, and interpretation.

Hyperoxia enhances slow-wave forebrain states in urethane-anesthetized and naturally sleeping rats.

Oxygen (O2) is a crucial element for physiological functioning in mammals. In particular, brain function is critically dependent on a minimum amount of circulating blood levels of O2 and both immediate and lasting neural dysfunction can result following anoxic or hypoxic episodes. Although the effects of deficiencies in O2 levels on the brain have been reasonably well studied, less is known about the influence of elevated levels of O2 (hyperoxia) in inspired gas under atmospheric pressure. This is of importance due to its typical use in surgical anesthesia, in the treatment of stroke and traumatic brain injury, and even in its recreational or alternative therapeutic use. Using local field potential (EEG) recordings in spontaneously breathing urethane-anesthetized and naturally sleeping rats, we characterized the influence of different levels of O2 in inspired gases on brain states. While rats were under urethane anesthesia, administration of 100% O2 elicited a significant and reversible increase in time spent in the deactivated (i.e., slow-wave) state, with concomitant decreases in both heartbeat and respiration rates. Increasing the concentration of carbon dioxide (to 5%) in inspired gas produced the opposite result on EEG states, mainly a decrease in the time spent in the deactivated state. Consistent with this, decreasing concentrations of O2 (to 15%) in inspired gases decreased time spent in the deactivated state. Further confirmation of the hyperoxic effect was found in naturally sleeping animals where it similarly increased time spent in slow-wave (nonrapid eye movement) states. Thus alterations of O2 in inspired air appear to directly affect forebrain EEG states, which has implications for brain function, as well as for the regulation of brain states and levels of forebrain arousal during sleep in both normal and pathological conditions. NEW & NOTEWORTHY We show that alterations of oxygen concentration in inspired air biases forebrain EEG state. Hyperoxia increases the prevalence of slow-wave states. Hypoxia and hypercapnia appear to do the opposite. This suggests that oxidative metabolism is an important stimulant for brain state.