Urethane Anesthesia Research Articles

Abstract WP234: Age And Type 2 Diabetes Reduce The Coherence Of Theta And Gamma Oscillations Between Brain Networks

Introduction: The integrity of the functional connectivity across networks relies on synchronous activation of neurons, which reduces with age. Type 2 diabetes (T2DM) is a major risk factor for stroke and cognitive decline and also a precocious aging, yet the electrophysiological changes in T2DM and its interaction with aging is not well understood. The current study sought to delineate the age and diabetic effect on the resting state field potential in regions crucial for cognition. Methods: Local field potential was recorded under urethane anesthesia with multichannel electrodes spanning the cortex and hippocampus (HPC) in control (db/+) and diabetic (db/db) mice, at 200 or 400 days of age. Brain state and neural activity in signal power of δ, θ, γ waves and sharp-wave associated ripples (SWRs) were determined. Coherence and phase-amplitude coupling (PAC) within the HPC or between cortex and HPC were computed to reflect the level of functional communication across brain networks. Results: Our results show that age decreased the θ/δ amplitude (T/D) ratio and the signal power of the theta, gamma and high gamma in the cortex and the HPC. Both age and diabetes prolonged the average duration of SWR clusters and inter-ripple intervals. The greatest SWR signal power was detected in db/db mice at 200 ds of age compared to the other 3 groups, so were the γ power in the cortex and HPC during SWRs. Both age and diabetes reduced θ coherence between cortex and HPC. Age also reduced γ coherence within the hippocampus. Interestingly, while age reduced the coherence of high γ band between hippocampus and CTX, diabetes significantly increased it. To the contrary, age increased coherence in δ within the hippocampal layers. Diabetes decreased the θ-low γ PAC between CA1 and cortex and within CA1. However, diabetes increased δ-γ PAC within the slm, while both age and diabetes increased the δ-γ PAC between the hippocampus and the cortex. Conclusion: The neural activity and synchrony in brain regions crucial for cognitive function is impaired by age and further confounded by T2DM. The perturbation of resting state field potential signal power and coherence may underlie age and diabetes induced loss of functional connectivity and cognitive decline.

Prefrontal-Hippocampal Pathways Through the Nucleus Reuniens Are Functionally Biased by Brain State.

Circuit-level communication between disparate brain regions is fundamental for the complexities of the central nervous system operation. Co-ordinated bouts of rhythmic activity between the prefrontal cortex (PFC) and hippocampus (HPC), in particular, are important for mnemonic processes. This is true during awake behavior, as well as during offline states like sleep. We have recently shown that the anatomically interposed thalamic nucleus reuniens (RE) has a role in coordinating slow-wave activity between the PFC and HPC. Here, we took advantage of spontaneous brain state changes occurring during urethane anesthesia in order to assess if PFC-HPC communication was modified during activated (theta) vs. deactivated (slow oscillation: SO) states. These forebrain states are highly similar to those expressed during rapid eye movement (REM) and non-REM stages of natural sleep, respectively. Evoked potentials and excitatory current sinks in the HPC were consistently larger during SO states, regardless of whether PFC or RE afferents were stimulated. Interestingly, PFC stimulation during theta appeared to preferentially use a cortico-cortical pathway, presumably involving the entorhinal cortex as opposed to the more direct RE to HPC conduit. Optogenetic and chemogenetic manipulations of the RE suggested that this state-dependent biasing was mediated by responding in the RE itself. Finally, the phase of both ongoing rhythms also appeared to be an important factor in modulating HPC responses, with maximal field excitatory postsynaptic potentials (EPSPs) occurring during the negative-going phase of both rhythms. Thus, forebrain state plays an important role in how communication takes place across the PFC and HPC, with the RE as a determining factor in how this is shaped. Furthermore, ongoing sleep-like rhythms influence the coordination and perhaps potentiate excitatory processing in this extended episodic memory circuit. Our results have direct implications for activity-dependent processes relevant to sleep-dependent memory consolidation.

Loss of Awareness or Urinary Dysfunction? Investigating Amyloidosis and Urinary Physiology in a Transgenic Mouse

Alzheimer’s disease (AD) is a devastating disorder primarily affecting older adults and is the most common neurodegenerative disease in the US. More than one in three AD patients experience AD-associated urinary dysfunction (ADUD), which directly contributes to their institutionalization. While ADUD has been clinically regarded as a result of poor cognitive control over urinary function, the physiology underlying loss of urinary control remains unknown. We hypothesize that amyloidosis in the CNS results in pathologic changes in urinary structure and function. Tg-APP/PS1DE9 mice were used before plaque deposition (4-6 months) and after plaque accumulation (8-10 months) and compared to WT littermates. Behavioral assays (open field testing and voiding spot assays) were performed to assess cortical function. Pressure-flow cystometry was conducted under urethane anesthesia to assess autonomic control of urinary function without cortical influence. Pharmacomyography of bladder strips was used to determine tissue-level changes in the absence of CNS input. In Tg-APP/PS1DE9 mice, plaque accumulation resulted in significant cystometric changes to voiding phase parameters, but not storage phase parameters. Pharmacologic studies showed decreased sensitivity to adrenergic stimulation without change in muscarinic sensitivity. Behavioral assays demonstrated significant differences between transgenic animals and WT in locomotion and voiding spot sizes. We interpret our data to support AD-related pathology of Aβ accumulation results in a distinct urinary phenotype in our model, analogous to the ADUD observed in AD patients. Establishing and verifying models of ADUD may improve the efficacy of treating ADUD and increase quality of life for patients and their caregivers.

Early-stage hippocampal hyperexcitability and impaired synaptic plasticity in the TgF344-AD rat model of Alzheimer's disease.

The transgenic TgF344-AD (TG) rat is emerging as a useful model for investigating age-dependent AD features that include behavioural, electrophysiological and pathological phenotypes of relevance to disease. In this study we investigated changes related to early stages of AD in 9 month old animals. Male and female TG rats and wild-type (WT) littermates undertook longitudinal behavioural testing at 3, 6 & 9 months (n=6-13/sex-genotype combination) and were tested for: (i) hippocampus-dependent rapid place learning in a delayed-matching-to place watermaze (DMP) test, (ii) novel object recognition (NOR) memory, (iii) expression of startle/pre-pulse inhibition; and (iv) open-field locomotor activity. In another cohort of rats at 9 months (n=8 males per genotype), spontaneous and evoked activities were record and measured in hippocampal CA3->CA1 circuit to study synaptic plasticity using in vivo electrophysiology under urethane anaesthesia. Following behavioural or electrophysiology testing brains were collected for post-mortem analyses (n=10 males per genotype). GABAergic and synaptic markers including parvalbumin, GAD67 , GABAA α5, SNAP-25 and PSD95 were measured by Simple Western analysis in the frontal cortex, prefrontal cortex, dorsal and ventral hippocampus. Behaviour: WT rats showed poor DMP performance with no clear genotype effect. Object exploration was low with no significant NOR. We found startle and locomotor sex differences, replicating previous reports. Interestingly, at 9 months, TG rats showed locomotor hypoactivity at the beginning and hyperactivity at the end of the 30-min open-field test, indicating impaired habituation, which may reflect impaired hippocampal contextual memory. Electrophysiology: 9-month-old TG rats showed reduced long-term potentiation with an increased baseline connectivity. A general decrease in power across frequency bands was found in the TG, especially for theta band. Western analysis: TG rats showed reduced levels of parvalbumin with a clear effect of genotype at 9-months. There was no detectable changes in synaptic markers. Impaired locomotor habituation of TG rats suggests abnormal hippocampal function emerges around 9 months, reflected by impaired hippocampal CA1 plasticity. A reduction of parvalbumin was also seen in the TG animals at this age. However, poor memory performance may limit the suitability of the TG model for testing clinically relevant memory deficits.

Acute treatment with dexketoprofen reduces penicillin induced epileptiform activity in wistar albino rats (dexketoprofen in penicillin induced seizure model)

AimDexketoprofen trometamol is one of the most commonly used anti-inflammatory analgesic agents for pain control. This study aims to investigate the effect of dexketoprofen on penicillin-induced epileptiform activity in rats. MethodIn this study, 28 male Wistar rats weighing 220–240 g were used. Tripolar electrodes were implanted under urethane anesthesia. Epileptiform activity was induced by micro-injection of 500 units (IU) penicillin into the rats’ left somatomotor cortex. Dexketoprofen (5, 25, and 50 mg/kg) was administrated intraperitoneally after 30 min of penicillin injection. Epileptiform activity was evaluated by electrocorticography (ECoG). ResultsThe low dose of dexketoprofen administration (5 mg/kg) reduced the mean spike frequency of epileptiform activity 60 min after its injection. However, 25 and 50 mg/kg dexketoprofen significantly reduced the mean spike frequency 30 min after the dexketoprofen injection compared to the control group (p < 0.05). The amplitudes of epileptiform discharges in all groups were unaffected (p > 0.05). ConclusionThis study revealed that dexketoprofen had a significant anti-seizure effect when applied at 5 mg/kg, 25 mg/kg, and 50 mg/kg (especially at 25 and 50 mg/kg), in the penicillin-induced seizure model. The obtained data revealed that dexketoprofen might play an essential role against epileptic seizures.

Long-term stability of physiological signals within fluctuations of brain state under urethane anesthesia.

Urethane, an acute laboratory anesthetic, produces distinct neurophysiological and physiological effects creating an effective model of the dynamics of natural sleep. As a model of both sleep-like neurophysiological activity and the downstream peripheral function urethane is used to model a variety of physiological and pathophysiological processes. As urethane is typically administered as a single-bolus dose, it is unclear the stability of peripheral physiological functions both within and between brain-states under urethane anesthesia. In this present study, we recorded respiration rate and heart rate concurrently with local field potentials from the neocortex and hippocampus to determine the stability of peripheral physiological functions within and between brain-states under urethane anesthesia. Our data shows electroencephalographic characteristics and breathing rate are remarkable stable over long-term recordings within minor reductions in heart rate on the same time scale. Our findings indicate that the use of urethane to model peripheral physiological functions associated with changing brain states are stable during long duration experiments.

Metabolism of hyperpolarised [1-13 C]pyruvate in awake and anaesthetised rat brains.

The use of hyperpolarised 13 C pyruvate for nononcological neurological applications has not been widespread so far, possibly due to delivery issues limiting the visibility of metabolites. First proof-of-concept results have indicated that metabolism can be detected in human brain, and this may supersede the results obtained in preclinical settings. One major difference between the experimental setups is that preclinical MRI/MRS routinely uses anaesthesia, which alters both haemodynamics and metabolism. Here, we used hyperpolarised [1-13 C]pyruvate to compare brain metabolism in awake rats and under isoflurane, urethane or medetomidine anaesthesia. Spectroscopic [1-13 C]pyruvate time courses measured sequentially showed that pyruvate-to-bicarbonate and pyruvate-to-lactate labelling rates were lower in isoflurane animals than awake animals. An increased bicarbonate-to-lactate ratio was observed in the medetomidine group compared with other groups. The study shows that hyperpolarised [1-13 C]pyruvate experiments can be performed in awake rats, thus avoiding anaesthesia-related issues. The results suggest that haemodynamics probably dominate the observed pyruvate-to-metabolite labelling rates and area-under-time course ratios of referenced to pyruvate. On the other hand, the results obtained with medetomidine suggest that the ratios are also modulated by the underlying cerebral metabolism. However, the ratios between intracellular metabolites were unchanged in awake compared with isoflurane-anaesthetised rats.

Recurrent febrile seizures alter intrahippocampal temporal coordination but do not cause spatial learning impairments.

Febrile seizures (FSs) are the most common form of seizures in children. Single short FSs are benign, but FSs lasting longer than 30min, termed febrile status epilepticus, may result in neurological sequelae. However, there is little information about an intermediary condition, brief recurrent FSs (RFSs). The goal of this study was to determine the role of RFSs on spatial learning and memory and the properties of spontaneous hippocampal signals. A hippocampus-dependent active avoidance task was used to assess spatial learning and memory in adult rats that underwent experimental RFSs (eRFSs) in early life compared with their littermate controls. Following completion of the task, we utilized high-density laminar probes to measure spontaneous hippocampal CA1 circuit activity under urethane anesthesia, which allowed for the simultaneous recording of input regions in CA1 associated with both CA3 and entorhinal cortex. RFSs did not result in deficits in the active avoidance spatial test, a hippocampus-dependent test of spatial learning and memory. However, in vivo high-density laminar electrode recordings from eRFS rats had significantly altered power and frequency expression of theta and gamma bandwidths as well as signaling efficacy along the CA1somatodendritic axis. Thus, although eRFS modified CA1 neuronal input/output dynamics, these alterations were not sufficient to impair active avoidance spatial behavior. These findings indicate that although eRFSs do not result in spatial cognitive deficits in the active avoidance task, recurrent seizures do alter the brain and result in longstanding changes in the temporal organization of the hippocampus.

Dysfunctional voiding behavior and impaired muscle contractility in a rat model of detrusor underactivity.

Detrusor underactivity (DU) is an understudied health concern with inadequate clinical management. The pathophysiology of DU is unclear, and current therapies fail to improve symptoms. The current studies characterized voiding function and contractility of bladder and urethral tissues in a novel rat model of DU. Female obese prone (OP) and obese resistant (OR) rats were fed a 60 kcal% fat diet at 8 weeks old. A subset of rats (n = 4/strain) underwent uroflowmetry biweekly for 18 weeks in metabolic cages. At 40-56 weeks old, rats (n = 9-10/strain) underwent instrumented cystometry under urethane anesthesia. Following cystometry, bladder and urethral tissues (n = 8-9/strain) were harvested for in vitro assessments of contractility in response to carbachol, electric field stimulation, atropine, alpha, beta-methylene ATP, and caffeine. OP rats exhibited increased urinary frequency (p = 0.0031), decreased voided volume (p = 0.0093), and urine flow rate (p = 0.0064) compared to OR rats during uroflowmetry. Bethanechol (10 mg/kg) did not alter uroflowmetry parameters. During cystometry, OP rats exhibited decreased bladder emptying efficiency (p < 0.0001), decreased pressure to generate a void (p < 0.0001), and increased EUS activity during filling (p = 0.0011). Bladder contractility was decreased in OP rats when exposed to carbachol (p < 0.0003) and ATP (p = 0.0004), whereas middle urethral contractility was increased when exposed to carbachol (p = 0.0014), EFS (p = 0.0289), and caffeine (p = 0.0031). Impaired cholinergic and purinergic signaling in the bladder may contribute to poor voiding function in OP rats. In addition, increased urethral activity may engage a guarding reflex to augment continence and exacerbate incomplete emptying.

Respiratory coupling between prefrontal cortex and hippocampus of rats anaesthetized with urethane in theta and non-theta states.

Respiratory modulation of forebrain activity, long considered hard to reliably separate from breathing artefacts, has been firmly established in recent years using a variety of advanced techniques. Respiratory-related oscillation (RRO) is derived from rhythmic nasal airflow in the olfactory bulb (OB) and is conveyed to higher order brain networks, including the prefrontal cortex (PFC) and hippocampus (HC), where it may potentially contribute to communication between these structures by synchronizing their activities at the respiratory rate. RRO was shown to change with sleep-wake states; it is strongest in quiet waking, somewhat less in active waking, characterized with theta activity in the HC, and absent in sleep. The goal of this study was to test RRO synchronization between PFC and HC under urethane anaesthesia where theta and non-theta states spontaneously alternate. We found that in theta states, PFC-HC coherences significantly correlated with OB-HC but not with OB-PFC, even though RRO was stronger in PFC than in HC. In non-theta states, PFC-HC synchrony correlated with coherences connecting OB to either PFC or HC. Thus, similar to freely behaving rats, PFC-HC synchrony at RRO was primarily dependent on the response of HC to the common rhythmic drive, but only in theta state. The findings help outlining the value and the limits of applications in which urethane-anaesthetized rats can be used for modelling the neural mechanisms of RRO in behaving animals.

Local field potentials identify features of cortico-hippocampal communication impacted by stroke and environmental enrichment therapy

Objective. Cognitive and memory impairments are common sequelae after stroke, yet how middle cerebral artery (MCA) stroke chronically affects the neural activity of the hippocampus, a brain region critical for memory but remote from the stroke epicenter, is poorly understood. Environmental enrichment (EE) improves cognition following stroke; however, the electrophysiology that underlies this behavioral intervention is still elusive. Approach. We recorded extracellular local field potentials simultaneously from sensorimotor cortex and hippocampus in rats during urethane anesthesia following MCA occlusion and subsequent EE treatment. Main results. We found that MCA stroke significantly impacted the electrophysiology in the hippocampus, in particular it disrupted characteristics of sharp-wave associated ripples (SPW-Rs) altered brain state, and disrupted phase amplitude coupling (PAC) within the hippocampus and between the cortex and hippocampus. Importantly, we show that EE mitigates stroke-induced changes to SPW-R characteristics but does not restore hippocampal brain state or PAC. Significance. These results begin to uncover the complex interaction between cognitive deficit following stroke and EE treatment, providing a testbed to assess different strategies for therapeutics following stroke.

Utilizing multimodal imaging to visualize potential mechanism for sudden death in epilepsy

Sudden death in epilepsy or SUDEP is a fatal condition that accounts for more than 4000 deaths each year. Limited clinical and preclinical data on sudden death suggest critical contributions from autonomic, cardiac, and respiratory pathways. A potential mechanism for such sudden and severe cardiorespiratory dysregulation may be linked to acid reflux-induced laryngospasm. Here, we expand on our previous investigations and utilize a novel multimodal approach to provide visual evidence of acid reflux-initiated cardiorespiratory distress and subsequent sudden death in seizing rats.We used systemic kainic acid to acutely induce seizure activity in Long Evans rats, under urethane anesthesia. We recorded electroencephalography (EEG), electrocardiography (ECG), chest plethysmography, and esophageal pH signals through a multimodal recording platform, during simultaneous fast MRI scans of the rat stomach and esophagus. MRI images, in correlation with electrophysiology data were used to identify seizure progression, stomach acid movement up the esophagus, cardiorespiratory changes, and sudden death.In all cases of sudden death, esophageal pH recordings alongside MRI images visualized stomach acid movement up the esophagus. Severe cardiac (ST segment elevation), respiratory (intermittent apnea) and brain activity (EEG narrowing due to hypoxia) changes were observed only after acid reached larynx, which strongly suggested onset of laryngospasm following acid reflux. The complementary information coming from electrophysiology and fast MRI scans provided insight into the mechanism of esophageal reflux, laryngospasm, obstructive apnea, and subsequent sudden death in seizing animals. The results carry clinical significance as it outlines a potential mechanism that may be relevant to SUDEP in humans.

Carbonic anhydrase inhibitors suppress seizures in a rat model of birth asphyxia.

Seizures are common in neonates recovering from birth asphyxia but there is general consensus that current pharmacotherapy is suboptimal and that novel antiseizure drugs are needed. We recently showed in a rat model of birth asphyxia that seizures are triggered by the post-asphyxia recovery of brain pH. Here our aim was to investigate whether carbonic anhydrase inhibitors (CAIs), which induce systemic acidosis, block the post-asphyxia seizures. The CAIs acetazolamide (AZA), benzolamide (BZA), and ethoxzolamide (EZA) were administered intraperitoneally or intravenously to 11-day-old rats exposed to intermittent asphyxia (30min; three 7+3 min cycles of 9% and 5% O2 at 20% CO2 ). Electrode measurements of intracortical pH, Po2 , and local field potentials (LFPs) were made under urethane anesthesia. Convulsive seizures and blood acid-base parameters were examined in freely behaving animals. The three CAIs decreased brain pH by 0.14-0.17 pH units and suppressed electrographic post-asphyxia seizures. AZA, BZA, and EZA differ greatly in their lipid solubility (EZA > AZA > BZA) and pharmacokinetics. However, there were only minor differences in the delay (range 0.8-3.7min) from intraperitoneal application to their action on brain pH. The CAIs induced a modest post-asphyxia elevation of brain Po2 that had no effect on LFP activity. AZA was tested in freely behaving rats, in which it induced a respiratory acidosis and decreased the incidence of convulsive seizures from 9 of 20 to 2 of 17 animals. AZA, BZA, and EZA effectively block post-asphyxia seizures. Despite the differences in their pharmacokinetics, they had similar effects on brain pH, which indicates that their antiseizure mode of action was based on respiratory (hypercapnic) acidosis resulting from inhibition of blood-borne and extracellular vascular carbonic anhydrases. AZA has been used for several indications in neonates, suggesting that it can be safely repurposed for the treatment of neonatal seizures as an add-on to the current treatment regimen.

Anesthetic protocols for urodynamic studies of the lower urinary tract in small rodents-A systematic review.

Urodynamic studies in rats and mice are broadly used to examine pathomechnisms of disease and identify and test therapeutic targets. This review aims to highlight the effects of the anesthetics on the lower urinary tract function and seeks to identify protocols that allow recovery from anesthesia and repeated measurements while preserving the function which is being studied. All studies published in English language, which compared the data obtained under various types of anesthesia and the urodynamics performed in awake animals were included. It appears that urethane, an anesthetic recommended extensively for the investigation of lower urinary tract function, is appropriate for acute urodynamic studies only. Major advantages of urethane are its stability and ability to preserve the micturition reflex. Due to its toxicity and carcinogenicity, urethane anesthesia should not be used for recovery procedures. This review evaluated available alternatives including propofol, isoflurane and combinations of urethane, ketamine/xylazine, ketamine/medetomidine, and/or fentanyl/fluanisone/midazolam. Different effects have been demonstrated among these drugs on the urinary bladder, the urethral sphincter, as well as on their neuroregulation. The lowest incidence of adverse effects was observed with the use of a combination of ketamine and xylazine. Although the variations in the reviewed study protocols represent a limitation, we believe that this summary will help in standardizing and optimizing future experiments.

Age-associated bladder and urethral coordination impairment and changes in urethral oxidative stress in rats

AimsWe examined age-associated changes in bladder and urethral coordination involving the nitric oxide (NO)/soluble guanylyl cyclase (sGC) system, which induces urethral smooth muscle relaxation, and urethral ischemic/oxidative stress changes in rats. Main methodsSixteen female Sprague-Dawley rats were divided into young (3 months old) and middle-aged (12–15 months old) groups. Urethral activity was evaluated by simultaneously recording intravesical pressure under isovolumetric conditions and urethral perfusion pressure (UPP) under urethane anesthesia. Sodium nitroprusside (SNP, 0.1 mg/kg), an NO donor, and BAY 41-2272, a novel NO-independent stimulator of sGC (0.1 mg/kg), were administered intravenously to both groups. N-nitro-l-arginine methyl ester hydrochloride (l-NAME, 100 mg/kg) was also injected intravenously, to inhibit NO synthase activity in both groups. Staining for the ischemic marker, hypoxia-inducible factor-1α (HIF-1α), and the oxidative stress markers, 8-hydroxy-2′-deoxyguanosine (8-OHdG) and malondialdehyde (MDA), was performed on tissue sections of the urethra, in both groups. Key findingsBaseline UPP and UPP changes were significantly lower in middle-aged rats than in young rats. After administration of SNP and BAY 41-2272, baseline UPP and UPP nadir were significantly decreased in both groups. After administration of l-NAME, UPP change/bladder contraction amplitude in young rats was still lower than at baseline but was completely restored to control levels in middle-aged rats. Immunoreactivity of HIF-1α, 8-OHdG, and MDA was higher in middle-aged rats than in young rats. SignificanceAge-associated ischemic and oxidative stress in the urethra might be correlated with impairment of the NO/sGC system and with coordination of the bladder and urethra.

In vivo electrophysiological mechanisms underlying cervical epidural stimulation in adult rats.

To electrophysiologically determine the predominant neural structures activated with cervical epidural stimulation (ES), well-established electrophysiological protocols (single-pulse, paired-pulse and multiple frequency stimulation) were delivered at rest, during motor activity and under anaesthesia in adult rats. Cervical ES resulted in spinal evoked motor responses with three different waveforms - early response (ER), middle response (MR) and late response (LR). ERs remained unmodulated by repeated stimulation protocols. In contrast, MRs and LRs were modulated by repeated stimulation protocols and volitional motor activity. ERs are consequential to the direct activation of motor efferents; MRs are secondary to type-I sensory afferent activation and LRs result from the engagement of wider spinal interneuronal circuitry with potential influence from supraspinal pathways. Evidence from this work is fundamental in enhancing our understanding of cervical ES, and critical in refining the design of neuromodulation-based rehabilitative strategies and in the construction of neuroprosthetics. Epidural stimulation (ES) of the lumbar spinal cord has demonstrated significant improvements in various physiological functions after a traumatic spinal cord injury in humans. Electrophysiological evidence from rodent, human and computational studies collectively suggest that the functional recovery following lumbar ES is mediated via direct activation of sensory afferent fibres. However, the mechanisms underlying cervical ES have not been comprehensively studied, which greatly limits our understanding of its effectiveness in restoring upper limb function. In this work, we determined the predominant neural structures that are activated with cervical ES using in vivo cervical spinal evoked motor responses (SEMRs). Standard electrophysiological protocols (single-pulse, paired-pulse and multiple frequency stimulation) were implemented in 11 awake and anaesthetized rats in four experimental stages. Three distinct types of cervical SEMRs were identified based on latency of their appearance: early response (ER), middle response (MR) and late response (LR). ERs remained unmodulated by repeated stimulation protocols. MRs and LRs were modulated by repeated stimulation protocols and volitional motor activity. Except for LRs being completely abolished under urethane, ketamine or urethane anaesthesia did not affect the appearance of cervical SEMRs. Our data, backed by literature, suggest that ERs are secondary to the direct activation of motor efferents, MRs are elicited by activation of type-I sensory afferents and LRs result from the engagement of interneuronal circuitry with potential influence from supraspinal pathways. The gathered information paves the way to designing motor rehabilitation strategies that can utilize cervical ES to recover upper limb function following neurological deficits.

Augmented Respiratory-Sympathetic Coupling and Hemodynamic Response to Acute Mild Hypoxia in Female Rodents With Chronic Kidney Disease.

Carotid body feedback and hypoxia may serve to enhance respiratory–sympathetic nerve coupling (respSNA) and act as a driver of increased blood pressure. Using the Lewis polycystic kidney (LPK) rat model of chronic kidney disease, we examined respSNA in adult female rodents with CKD and their response to acute hypoxia or hypercapnia compared to Lewis control animals. Under urethane anesthesia, phrenic nerve activity, splanchnic sympathetic nerve activity (sSNA), and renal sympathetic nerve activity (rSNA) were recorded under baseline conditions and during mild hypoxic or hypercapnic challenges. At baseline, tonic SNA and blood pressure were greater in female LPK rats versus Lewis rats (all P < 0.05) and respSNA was at least two-fold larger [area under the curve (AUC), sSNA: 7.8 ± 1.1 vs. 3.4 ± 0.7 μV s, rSNA: 11.5 ± 3 vs. 4.8 ± 0.7 μV s, LPK vs. Lewis, both P < 0.05]. Mild hypoxia produced a larger pressure response in LPK [Δ mean arterial pressure (MAP) 30 ± 6 vs. 12 ± 6 mmHg] and augmented respSNA (ΔAUC, sSNA: 8.9 ± 3.4 vs. 2 ± 0.7 μV s, rSNA: 6.1 ± 1.2 vs. 3.1 ± 0.7 μV s, LPK vs. Lewis, all P ≤ 0.05). In contrast, central chemoreceptor stimulation produced comparable changes in blood pressure and respSNA (ΔMAP 13 ± 3 vs. 9 ± 5 mmHg; respSNA ΔAUC, sSNA: 2.5 ± 1 vs. 1.3 ± 0.7 μV s, rSNA: 4.2 ± 0.9 vs. 3.5 ± 1.4 μV s, LPK vs. Lewis, all P > 0.05). These results demonstrate that female rats with CKD exhibit heightened respSNA coupling at baseline that is further augmented by mild hypoxia, and not by hypercapnia. This mechanism may be a contributing driver of hypertension in this animal model of CKD.

Aloin attenuates chronic constriction injury-induced neuropathic pain in rats by inhibiting inflammatory cytokines and oxidative stress

Objective: To investigate the effect of aloin against chronic constriction injury (CCI)-induced neuropathic pain in rats. Methods: Rats were randomly divided into 7 groups: Group I (normal control), Group II (sham-operated), Group III (CCI control) and Group IV, V, VI, and VII, which underwent CCI surgery and then were administered with aloin (5 mg/kg, p.o.; 25 mg/kg, p.o.; 125 mg/kg, p.o.) and gabapentin (50 mg/kg, p.o.), respectively for 14 days. Peripheral neuropathy was induced by silk ligatures (4-0) loosely placed around the sciatic nerve. Nociceptive thresholds against mechanical stimuli (Von-Frey filaments) and thermal stimuli (12 °C and 40 °C) were measured at mid-plantar paw region ipsilateral to the compressed nerve on day-3, 7, 11, and 14. The concentration of cytokines including tumor necrosis factor-α (TNF-α), interleukin-6, and interleukin-1β was estimated at day-7. At day 14, motor nerve conduction velocity was determined under urethane anesthesia (1.25 g/kg). Oxidative stress parameters (malondiadehyde, glutathione, catalase, and superoxide dismutase) were estimated in sciatic nerve homogenates at day 14. Representative nerve samples were processed for histological investigations. Results: Aloin significantly reduced CCI-induced mechanical and thermal allodynia. It also improved motor nerve conduction velocity and decreased oxidative stress in nerve tissues. In addition, it decreased pro-inflammatory cytokine levels and restored the histoarchitecture of compressed sciatic nerve. Conclusions: Aloin mitigates CCI-induced neuropathic pain in rats by inhibiting oxidative stress and pro-inflammatory cytokines in the afflicted sciatic nerve.

Sympathetic responses during renal artery occlusion are altered by anesthesia

Sympathetic hyperactivity contributes to stenotic renovascular hypertension. Transient or prolonged reduction in renal perfusion or renal artery occlusion activates renal sensory fibers to alter renal hemodynamics. Electrical stimulation of these fibers increases sympathetic nerve activity (SNA) and arterial blood pressure (ABP). Recently, we demonstrated that anesthetic agents differentially impact renal afferent sensitivity to mechano- and/or chemical-stimuli, including acute renal artery occlusion. The purpose of the present study was to assess the extent by which anesthesia affects efferent sympathetic reflex responses to renal artery occlusion. Renal, splanchnic, and lumbar SNA were measured during transient renal artery occlusion (3 min) in male and female Sprague-Dawley rats (250-400g) using unanesthetized decerebrate preparations, Inactin, isoflurane, or urethane anesthesia. Baseline ABP was significantly higher for decerebrate (122±8 mmHg) and urethane (133±6 mmHg) groups versus Inactin (78±3 mmHg) and isoflurane (95±4 mmHg; p=0.0001). Within the initial 30 seconds, renal artery occlusion decreased mean ABP in decerebrate (Δmean ABP= -13±3 mmHg; n=8; 5M, 3F), Inactin (Δmean ABP= -7±1 mmHg; n=11; 6M, 5F), urethane (Δmean ABP= -19±4 mmHg; n=11; 6M, 5F), and Isoflurane (Δmean ABP= -6±3 mmHg; n=10; 5M, 5F) groups. SNA was differentially altered during renal artery occlusion. Renal SNA significantly increased in the decerebrate (149±24%), Inactin (130±9%), and urethane (144±24%) groups relative to baseline but was significantly reduced in the isoflurane group (79±2%). Splanchnic SNA was similarly increased in decerebrate (130±12%), Inactin (150±23%), and urethane (147±19%) groups yet was significantly reduced in the isoflurane group (79±4%) compared to baseline. In marked contrast, renal artery occlusion decreased lumbar SNA in all groups (Decerebrate: 68±7%; Inactin: 67±5%; urethane: 58±3%; and Isoflurane: 63±7%. In conclusion, anesthesia differentially influences SNA and ABP responses during renal artery occlusion. In the majority of groups, arterial occlusion a differential SNA response characterized by increased renal and splanchnic SNA but decreased lumbar SNA. Isoflurane anesthesia attenuated sympathoexcitation across all groups.

High Salt Diet Blunted Baroreflex Sensitivity (BRS) to Bilateral Common Carotid Artery Occlusion (BCCO) in Male Sprague –Dawley Rats – Permissive Effect of Testosterone

Autonomic dysfunction is involved in the onset of salt-sensitive hypertension as baroreflex sensitivity (BRS) is blunted in salt-sensitive hypertension. However, salt sensitivity exhibits sex disparity because in salt-sensitive hypertension, blood pressure (BP) is usually higher in males when compared with females. The mechanism(s) underlying this sexual dimorphism is not clear. We, therefore, designed this study to determine BRS in weanling male Sprague – Dawley rats that were either bilaterally orchidectomized or sham-operated (under ketamine and xylazine anaesthesia (90 mg and 10 mg/kg/body weight i.m) respectively, with or without testosterone replacement (10 mg/kg sustanon 250® i.m once in 3 weeks) and were placed on normal (0.3%) or high (8%) NaCl diet for 6 weeks. Blood pressure (BP) and heart rate (HR) were measured via arterial cannulation under urethane and α-chloralose anesthesia (5 ml/kg body weight i.p). BRS was determined as change in HR per unit change in BP in response to 30 seconds bilateral common carotid artery occlusion (BCCO). Serum concentration of testosterone was measured using enzyme-linked immunosorbent assay (ELISA). High salt diet (HSD) increased both the basal BP and peak BP after BCCO when compared with control. Orchidectomy attenuated but testosterone replacement restored the elevated BP in rats fed HSD. HSD blunted BRS to BCCO. However, this effect of HSD was lost in the absence of testosterone as there was no significant difference in BRS between rat fed a normal or high salt diet. Our results suggest that blunting of the BRS may be one of the mechanisms by which testosterone promotes salt-induced hypertension and serves as one of the bases for sex disparity in salt-sensitive hypertension.