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Cholinergic modulation inhibits cortical spreading depression in mouse neocortex through activation of muscarinic receptors and decreased excitatory/inhibitory drive.

Cortical spreading depression (CSD) is a wave of transient network hyperexcitability leading to long lasting depolarization and block of firing, which initiates focally and slowly propagates in the cerebral cortex. It causes migraine aura and it has been implicated in the generation of migraine headache. Cortical excitability can be modulated by cholinergic actions, leading in neocortical slices to the generation of rhythmic synchronous activities (UP/DOWN states). We investigated the effect of cholinergic activation with the cholinomimetic agonist carbachol on CSD triggered with 130 mM KCl pulse injections in acute mouse neocortical brain slices, hypothesizing that the cholinergic-induced increase of cortical network excitability during UP states could facilitate CSD. We observed instead an inhibitory effect of cholinergic activation on both initiation and propagation of CSD, through the action of muscarinic receptors. In fact, carbachol-induced CSD inhibition was blocked by atropine or by the preferential M1 muscarinic antagonist telenzepine; the preferential M1 muscarinic agonist McN-A-343 inhibited CSD similarly to carbachol, and its effect was blocked by telenzepine. Recordings of spontaneous excitatory and inhibitory post-synaptic currents in pyramidal neurons showed that McN-A-343 induced overall a decrease of the excitatory/inhibitory ratio. This inhibitory action may be targeted for novel pharmacological approaches in the treatment of migraine with muscarinic agonists.

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Aquaporin 4 knockout increases complete Freund’s adjuvant-induced spinal central sensitization.

Growing evidence suggests a critical role of astrocytes for pain regulation. The water channel protein aquaporin 4 (AQP4), a functional regulator of astrocytes, is involved in various neurological disorders. However, the pathophysiological roles of AQP4 in pain conditions remain unclear. In the present study, we investigated the effect of AQP4 gene knockout in central sensitization induced by complete Freund's adjuvant (CFA). The behavioral analysis revealed that mechanical allodynia and thermal hyperalgesia were more severe in AQP4 null mice than those of wild-type controls over the course of 11 days following CFA intraplantar injection. CFA caused activation of astrocytes with upregulated expression levels of AQP4 and glutamate transporter 1 (GLT1) in the dorsal horn of the spinal cord. AQP4 deficiency reduced GLT1 up-regulation, causing persistent expression of the neuronal activation marker Fos within superficial dorsal horn neurons, including glutamatergic neurons. However, AQP4 deletion did not affect CFA-evoked proinflammatory cytokine expression in the spinal cord. Together, these results have shown that AQP4 absence intensifies CFA-induced spinal central sensitization, which is associated with reduced compensatory up-regulation of GLT1, subsequently increasing glutamatergic overexcitation. Therefore, targeting spinal cord AQP4 may serve as a potential strategy for treatment of peripheral inflammation-evoked hyperalgesia.

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Attenuated dopamine receptor signaling in nucleus accumbens core in a rat model of chemically-induced neuropathy.

Neuropathy is major source of chronic pain that can be caused by mechanically or chemically induced nerve injury. Intraplantar formalin injection produces local necrosis over a two-week period and has been used to model neuropathy in rats. To determine whether neuropathy alters dopamine (DA) receptor responsiveness in mesolimbic brain regions, we examined dopamine D-like and D-like receptor (DR) signaling and expression in male rats 14 days after bilateral intraplantar formalin injections into both rear paws. DR-mediated G-protein activation and expression of the DR long, but not short, isoform were reduced in nucleus accumbens (NAc) core, but not in NAc shell, caudate-putamen or ventral tegmental area of formalin-compared to saline-treated rats. In addition, DR-stimulated adenylyl cyclase activity was also reduced in NAc core, but not in NAc shell or prefrontal cortex, of formalin-treated rats, whereas DR expression was unaffected. Other proteins involved in dopamine neurotransmission, including dopamine uptake transporter and tyrosine hydroxylase, were unaffected by formalin treatment. In behavioral tests, the potency of a DR agonist to suppress intracranial self-stimulation (ICSS) was decreased in formalin-treated rats, whereas DR agonist effects were not altered. The combination of reduced DR expression and signaling in NAc core with reduced suppression of ICSS responding by a DR agonist suggest a reduction in D autoreceptor function. Altogether, these results indicate that intraplantar formalin produces attenuation of highly specific DA receptor signaling processes in NAc core of male rats and suggest the development of a neuropathy-induced allostatic state in both pre- and post-synaptic DA receptor function.

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Celecoxib reduces CSD-induced macrophage activation and dilatation of dural but not pial arteries in rodents: implications for mechanism of action in terminating migraine attacks.

Non-steroidal anti-inflammatory drugs (NSAIDs), commonly known as COX-1/COX-2 inhibitors, can be effective in treating mild to moderate migraine headache. However, the mechanism by which these drugs act in migraine is not known, nor is the specific contribution of COX-1 versus COX-2 known. We sought to investigate these unknowns using celecoxib, which selectively inhibits the enzymatic activity of COX-2, by determining its effects on several migraine-associated vascular and inflammatory events. Using in vivo two-photon microscopy, we determined intraperitoneal celecoxib effects on CSD-induced blood vessel responses, plasma protein extravasation, and immune cell activation in the dura and pia of mice and rats. Compared to vehicle (control group), celecoxib reduced significantly CSD-induced dilatation of dural arteries and activation of dural and pial macrophages but not dilatation or constriction of pial arteries and veins, or the occurrence of plasma protein extravasation. Collectively, these findings suggest that a mechanism by which celecoxib-mediated COX-2 inhibition might ease the intensity of migraine headache and potentially terminate an attack is by attenuating dural macrophages activation and arterial dilatation outside the blood brain barrier (BBB), and pial macrophages activation inside the BBB.

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Neurons differentiate magnitude and location of mechanical stimuli.

Neuronal activity can be modulated by mechanical stimuli. To study this phenomenon quantitatively, we mechanically stimulated rat cortical neurons by shear stress and local indentation. Neurons show 2 distinct responses, classified as transient and sustained. Transient responses display fast kinetics, similar to spontaneous neuronal activity, whereas sustained responses last several minutes before returning to baseline. Local soma stimulations with micrometer-sized beads evoke transient responses at low forces of ∼220 nN and pressures of ∼5.6 kPa and sustained responses at higher forces of ∼360 nN and pressures of ∼9.2 kPa. Among the neuronal compartments, axons are highly susceptible to mechanical stimulation and predominantly show sustained responses, whereas the less susceptible dendrites predominantly respond transiently. Chemical perturbation experiments suggest that mechanically evoked responses require the influx of extracellular calcium through ion channels. We propose that subtraumatic forces/pressures applied to neurons evoke neuronal responses via nonspecific gating of ion channels.

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The sad weekend: A perilous North American tradition.

Obesity is a global concern and affects millions of Americans who consume poor-quality diets. Diets directly affect the gut microbiota, which can have subsequent effects on inflammation and contribute to other chronic states. Previously we have shown that a Standard American Diet (SAD) increased immune cell activation and prolonged recovery and that a beneficial diet could reduce these negative effects. Here, male and female mice were given access to regular chow (REG), SAD, our Anti-Inflammatory Diet (AID) or a combination of SAD and AID. This latter group was modeled on the commonplace dietary pattern of healthy eating during the week (AID: Monday-Friday) and relaxed eating patterns on the weekend (SAD: Saturday-Sunday). After 14 weeks of diet consumption and an inflammatory injury, we found that the SAD prolonged and the AID promoted recovery. However, recovery was significantly delayed in those mice consuming the AID-SAD, regardless of weekly healthy diet access. In addition, fecal samples taken during the study revealed dramatic differences in microbial community composition, relative abundance of abundant bacterial phyla and alpha diversity. These data confirm the impact of diet on gut microbiota and suggest a relation between abundance of specific bacterial taxa and susceptibility to prolonged recovery from injury.

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Chronic inflammatory pain alters alcohol-regulated frontocortical signaling and associations between alcohol drinking and thermal sensitivity.

Alcohol use disorder (AUD) is a chronic, relapsing psychiatric disorder that is characterized by the emergence of negative affective states. The transition from recreational, limited intake to uncontrolled, escalated intake is proposed to involve a transition from positive to negative reinforcement mechanisms for seeking alcohol. Past work has identified the emergence of significant hyperalgesia/allodynia in alcohol-dependent animals, which may serve as a key negative reinforcement mechanism. Chronic pain has been associated with enhanced extracellular signal-regulated kinase (ERK) activity in cortical and subcortical nociceptive areas. Additionally, both pain and AUD have been associated with increased activity of the glucocorticoid receptor (GR), a key mediator of stress responsiveness. The objectives of the current study were to first determine relationships between thermal nociceptive sensitivity and alcohol drinking in male Wistar rats. While inflammatory pain induced by complete Freund's adjuvant (CFA) administration did not modify escalation of home cage drinking in animals over four weeks, the relationship between drinking levels and hyperalgesia symptoms reversed between acute (1 week) and chronic (3-4 week) periods post-CFA administration, suggesting that either the motivational or analgesic effects of alcohol may be altered over the time course of chronic pain. We next examined ERK and GR phosphorylation in pain-related brain areas (including the central amygdala and prefrontal cortex subregions) in animals experiencing acute withdrawal from binge alcohol administration (2 g/kg, 6 h withdrawal) and CFA administration (four weeks) to model the neurobiological consequences of binge alcohol exposure in the context of pain. We observed a significant interaction between alcohol and pain state, whereby alcohol withdrawal increased ERK phosphorylation across all four frontocortical areas examined, although this effect was absent in animals experiencing chronic inflammatory pain. Alcohol withdrawal also increased GR phosphorylation across all four frontocortical areas, but these changes were not altered by CFA. Interestingly, we observed significant inter-brain regional correlations in GR phosphorylation between the insula and other regions investigated only in animals exposed to both alcohol and CFA, suggesting coordinated activity in insula circuitry and glucocorticoid signaling in this context. The results of these studies provide a greater understanding of the neurobiology of AUD and will contribute to the development of effective treatment strategies for comorbid AUD and pain.

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Microglia-mediated chronic psoriatic itch induced by imiquimod.

Activation of glial cells has been shown to play an important role in chronic itch. However, whether glial cells play an important role in the development of psoriasis-induced chronic itch has not been fully elucidated. This study investigated the role of spinal glial cells in psoriasis-induced chronic itch. To develop a mouse model of psoriasis-induce chronic itch, we used 5% imiquimod cream to receive a daily topical application on the shaved back skin for seven consecutive days. The results showed that the expression of microglial marker ionized calcium binding adaptor molecule-1 was significantly increased after 5% imiquimod treatment in cervical spinal cord dorsal horn (C3-C4), and the intrathecal microglial inhibitor minocycline or PLX5622 diet suppressed both spontaneous itch and microglial activation. Furthermore, we found that the number of scratches and alloknesis score in female mice was significantly greater than in male mice after 5% imiquimod treatment. Our results indicate that microglia mediate chronic psoriatic itch induced by imiquimod.

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Longitudinal FDG-PET scan study of brain changes in mice with cancer-induced bone pain and after morphine analgesia.

Morphine is the most commonly used drug for treating physical and psychological suffering caused by advanced cancer. Although morphine is known to elicit multiple supraspinal analgesic effects, its behavioral correlates with respect to the whole brain metabolic activity during cancer-induced bone pain have not been elucidated. We injected 4T1 mouse breast cancer cells into the left femur bone marrow cavity of BALB/c mice. All mice developed limb use deficits, mechanical allodynia, and hypersensitivity to cold, which were effectively suppressed with morphine. Serial 18F-fluorodeoxyglucose positron emission tomography (FDG-PET) was performed for each mouse before cancer induction (0 d), after cancer-induced bone pain was established (14 d), and during effective morphine treatment (16 d). The longitudinal FDG-PET imaging analysis demonstrated that cancer-induced bone pain increased glucose uptake in the insular cortex and hypothalamus and decreased the activity of the retrosplenial cortex. Morphine reversed the activation of the insular cortex and hypothalamus. Furthermore, morphine activated the amygdala and rostral ventromedial medulla and suppressed the activity of anterior cingulate cortex. Our findings of hypothalamic and insular cortical activation support the hypothesis that cancer-induced bone pain has strong inflammatory and affective components in freely moving animals. Morphine may provide descending inhibitory and facilitatory actions in the treatment of cancer-induced bone pain in a clinical setting.

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Altered glial glutamate transporter expression in descending circuitry and the emergence of pain chronicity.

The glutamate type 1 transporter (GLT1) plays a major role in glutamate homeostasis in the brain. Although alterations of GLT1 activity have been linked to persistent pain, the significance of these changes is poorly understood. Focusing on the rostral ventromedial medulla, a key site in pain modulation, we examined the expression and function of GLT1 and related transcription factor kappa B-motif binding phosphoprotein (KBBP) in rats after adjuvant-induced hind paw inflammation.

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