(-)-Phenserine and the prevention of pre-programmed cell death and neuroinflammation in mild traumatic brain injury and Alzheimer's disease challenged mice.

Mild traumatic brain injury (mTBI) is a risk factor for neurodegenerative disorders, such as Alzheimer's disease (AD) and Parkinson's disease (PD). TBI-derived neuropathologies are promoted by inflammatory processes: chronic microgliosis and release of pro-inflammatory cytokines that further promote neuronal dysfunction and loss. Herein, we evaluated the effect on pre-programmed cell death/neuroinflammation/synaptic integrity and function of (-)-Phenserine tartrate (Phen), an agent originally developed for AD. This was studied at two clinically translatable doses (2.5 and 5.0 mg/kg, BID), in a weight drop (concussive) mTBI model in wild type (WT) and AD APP/PSEN1 transgenic mice. Phen mitigated mTBI-induced cognitive impairment, assessed by Novel Object Recognition and Y-maze behavioral paradigms, in WT mice. Phen fully abated mTBI-induced neurodegeneration, evaluated by counting Fluoro-Jade C-positive (FJC+) cells, in hippocampus and cortex of WT mice. In APP/PSEN1 mice, degenerating cell counts were consistently greater across all experimental groups vs. WT mice. mTBI elevated FJC+ cell counts vs. the APP/PSEN1 control (sham) group, and Phen similarly mitigated this. Anti-inflammatory effects on microglial activation (IBA1-immunoreactivity (IR)) and the pro-inflammatory cytokine TNF-α were evaluated. mTBI increased IBA1-IR and TNF-α/IBA1 colocalization vs. sham, both in WT and APP/PSEN1 mice. Phen decreased IBA1-IR throughout hippocampi and cortices of WT mice, and in cortices of AD mice. Phen, likewise, reduced levels of IBA1/TNF-α-IR colocalization volume across all areas in WT animals, with a similar trend in APP/PSEN1 mice. Actions on astrocyte activation by mTBI were followed by evaluating GFAP, and were similarly mitigated by Phen. Synaptic density was evaluated by quantifying PSD-95+ dendritic spines and Synaptophysin (Syn)-IR. Both were significantly reduced in mTBI vs. sham in both WT and APP/PSEN1 mice. Phen fully reversed the PSD-95+ spine loss in WT and Syn-IR decrease in both WT and APP/PSEN1 mice. To associate immunohistochemical changes in synaptic markers with function, hippocampal long term potentiation (LTP) was induced in WT mice. LTP was impaired by mTBI, and this impairment was mitigated by Phen. In synopsis, clinically translatable doses of Phen ameliorated mTBI-mediated pre-programmed cell death/neuroinflammation/synaptic dysfunction in WT mice, consistent with fully mitigating mTBI-induced cognitive impairments. Phen additionally demonstrated positive actions in the more pathologic brain microenvironment of AD mice, further supporting consideration of its repurposing as a treatment for mTBI.

Pharmacokinetics and efficacy of PT302, a sustained-release Exenatide formulation, in a murine model of mild traumatic brain injury.

Traumatic brain injury (TBI) is a neurodegenerative disorder for which no effective pharmacological treatment is available. Glucagon-like peptide 1 (GLP-1) analogues such as Exenatide have previously demonstrated neurotrophic and neuroprotective effects in cellular and animal models of TBI. However, chronic or repeated administration was needed for efficacy. In this study, the pharmacokinetics and efficacy of PT302, a clinically available sustained-release Exenatide formulation (SR-Exenatide) were evaluated in a concussive mild (m)TBI mouse model. A single subcutaneous (s.c.) injection of PT302 (0.6, 0.12, and 0.024 mg/kg) was administered and plasma Exenatide concentrations were time-dependently measured over 3 weeks. An initial rapid regulated release of Exenatide in plasma was followed by a secondary phase of sustained-release in a dose-dependent manner. Short- and longer-term (7 and 30 day) cognitive impairments (visual and spatial deficits) induced by weight drop mTBI were mitigated by a single post-injury treatment with Exenatide delivered by s.c. injection of PT302 in clinically translatable doses. Immunohistochemical evaluation of neuronal cell death and inflammatory markers, likewise, cross-validated the neurotrophic and neuroprotective effects of SR-Exenatide in this mouse mTBI model. Exenatide central nervous system concentrations were 1.5% to 2.0% of concomitant plasma levels under steady-state conditions. These data demonstrate a positive beneficial action of PT302 in mTBI. This convenient single, sustained-release dosing regimen also has application for other neurological disorders, such as Alzheimer's disease, Parkinson's disease, multiple system atrophy and multiple sclerosis where prior preclinical studies, likewise, have demonstrated positive Exenatide actions.

Functional effects of synthetic cannabinoids versus Δ9 -THC in mice on body temperature, nociceptive threshold, anxiety, cognition, locomotor/exploratory parameters and depression.

Synthetic cannabinoids are psychoactive substances designed to mimic the euphorigenic effects of the natural cannabis. Novel unregulated compounds appear once older compounds become illegal. It has been previously reported that synthetic cannabinoids are different than Δ9 -tetrahydrocannabinol (Δ9 -THC) as they have chemical structures unrelated to Δ9 -THC, different metabolism and, often, greater toxicity. This study aimed to investigate the effects of three novel synthetic cannabinoids and pure Δ9 -THC on body temperature, nociceptive threshold, anxiety, memory function, locomotor and exploratory parameters, and depression. We performed a battery of behavioural and motor tests starting 50 minutes post i.p. injection of each drug to adult ICR mice. The synthetic cannabinoids that were used are AB-FUBINACA, AB-CHMINACA and PB-22. All synthetic cannabinoids and Δ9 -THC caused hypothermia, but only Δ9 -THC induced a clear antinociceptive effect. All synthetic cannabinoids and Δ9 -THC caused decreased anxiety levels, spatial memory deficits and decreased exploratory behaviour as measured in the elevated plus maze, Y-maze and staircase paradigm, respectively. However, all synthetic cannabinoids but not Δ9 -THC demonstrated decreased locomotor activity in the staircase test. Moreover, only AB-FUBINACA and Δ9 -THC affected the gait balance and grip strength of the mice as was assessed by the latency time to fall from a rod. In the forced swimming test, PB-22 caused elevated depression-like behaviour while AB-FUBINACA induced a reversed effect. These results suggest varied effects among different synthetic cannabinoids and Δ9 -THC. Further studies are needed to characterize the overall effects and differences between these synthetic cannabinoids and Δ9 -THC.

Liraglutide is neurotrophic and neuroprotective in neuronal cultures and mitigates mild traumatic brain injury in mice.

Traumatic brain injury (TBI), a brain dysfunction for which there is no present effective treatment, is often caused by a concussive impact to the head and affects an estimated 1.7 million Americans annually. Our laboratory previously demonstrated that exendin-4, a long-lasting glucagon-like peptide 1 receptor (GLP-1R) agonist, has neuroprotective effects in cellular and animal models of TBI. Here, we demonstrate neurotrophic and neuroprotective effects of a different GLP-1R agonist, liraglutide, in neuronal cultures and a mouse model of mild TBI (mTBI). Liraglutide promoted dose-dependent proliferation in SH-SY5Y cells and in a GLP-1R over-expressing cell line at reduced concentrations. Pre-treatment with liraglutide rescued neuronal cells from oxidative stress- and glutamate excitotoxicity-induced cell death. Liraglutide produced neurotrophic and neuroprotective effects similar to those of exendin-4 in vitro. The cAMP/PKA/pCREB pathway appears to play an important role in this neuroprotective activity of liraglutide. Furthermore, our findings in cell culture were well-translated in a weight drop mTBI mouse model. Post-treatment with a clinically relevant dose of liraglutide for 7 days in mice ameliorated memory impairments caused by mTBI when evaluated 7 and 30 days post trauma. These data cross-validate former studies of exendin-4 and suggest that liraglutide holds therapeutic potential for the treatment of mTBI. Exendin-4, a long-lasting glucagon-like peptide 1 receptor (GLP-1R) agonist, has neuroprotective effects in cellular and animal models of traumatic brain injury (TBI). Here, we demonstrate neurotrophic and neuroprotective effects of a different GLP-1R agonist, liraglutide, in neuronal cultures and a mouse model of mild TBI (mTBI). Liraglutide promoted dose-dependent proliferation in SH-SY5Y cells and in a GLP-1R over-expressing cell line at reduced concentrations. Pretreatment with liraglutide rescued neuronal cells from oxidative stress- and glutamate excitotoxicity-induced cell death. Liraglutide produced neurotrophic and neuroprotective effects similar to those of exendin-4 in vitro, likely involving the cAMP/PKA/pCREB pathway. Our findings in cell culture were well-translated in a weight-drop mTBI mouse model. Post-treatment with a clinically relevant dose of liraglutide for 7 days in mice ameliorated memory impairments caused by mTBI.

Sex-specific cognitive effects of mild traumatic brain injury to the frontal and temporal lobes.

BACKGROUND: Cognitive deficits are the most enduring and debilitating sequelae of mild traumatic brain injury (mTBI). However, relatively little is known about whether the cognitive effects of mTBI vary with respect to time post-injury, biological sex, and injury location. OBJECTIVES: The aim of this study was to assess the effect of the side and site of mTBI and to determine whether these effects are sexually dimorphic. METHODS: Male and female ICR mice were subjected to either a sham procedure or mTBI to the temporal lobes (right-sided or left-sided) or to the frontal lobes (bilateral) using a weight-drop model. After recovery, mice underwent a battery of behavioral tests at two post-injury time points. RESULTS: Different mTBI impact locations produced dissociable patterns of memory deficits; the extent of these deficits varied across sexes, time points, and memory domains. In both sexes, frontal mTBI mice exhibited a delayed onset of spatial memory deficits. Additionally, the performance of the frontal and left temporal injured males and females was more variable than that of controls. Interestingly, only in females does the effect of mTBI on visual recognition memory depend on the time post-injury. Moreover, only in females does spatial recognition memory remain relatively intact after mTBI to the left temporal lobe. CONCLUSION: This study showed that different mTBI impact sites produce dissociable and sex-specific patterns of cognitive deficits in mice. The results emphasize the importance of considering the injury site/side and biological sex when evaluating the cognitive sequelae of mTBI.

Author Correction: Bone Anabolic Response in the Calvaria Following Mild Traumatic Brain Injury is Mediated by the Cannabinoid-1 Receptor.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Bone Anabolic Response in the Calvaria Following Mild Traumatic Brain Injury is Mediated by the Cannabinoid-1 Receptor.

Brain trauma was clinically associated with increased osteogenesis in the appendicular skeleton. We showed previously in C57BL/6J mice that mild traumatic brain injury (mTBI) transiently induced bone formation in the femur via the cannabinoid-1 (CB1) receptor. Here, we subjected ICR mice to mTBI and examined the bone response in the skull using microCT. We also measured mast cell degranulation (MCD)72 h post-injury. Finally, we measured brain and calvarial endocannabinoids levels post-mTBI. mTBI led to decreased bone porosity on the contralateral (untouched) side. This effect was apparent both in young and mature mice. Administration of rimonabant (CB1 inverse agonist) completely abrogated the effect of mTBI on calvarial porosity and significantly reduced MCD, compared with vehicle-treated controls. We also found that mTBI resulted in elevated levels of anandamide, but not 2-arachidonoylglycerol, in the contralateral calvarial bone, whereas brain levels remained unchanged. In C57BL/6J CB1 knockout mice, mTBI did not reduce porosity but in general the porosity was significantly lower than in WT controls. Our findings suggest that mTBI induces a strain-specific CB1-dependent bone anabolic response in the skull, probably mediated by anandamide, but seemingly unrelated to inflammation. The endocannabinoid system is therefore a plausible target in management of bone response following head trauma.

Neuroprotective Effects and Treatment Potential of Incretin Mimetics in a Murine Model of Mild Traumatic Brain Injury.

Traumatic brain injury (TBI) is a commonly occurring injury in sports, victims of motor vehicle accidents, and falls. TBI has become a pressing public health concern with no specific therapeutic treatment. Mild TBI (mTBI), which accounts for approximately 90% of all TBI cases, may frequently lead to long-lasting cognitive, behavioral, and emotional impairments. The incretins glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) are gastrointestinal hormones that induce glucose-dependent insulin secretion, promote ß-cell proliferation, and enhance resistance to apoptosis. GLP-1 mimetics are marketed as treatments for type 2 diabetes mellitus (T2DM) and are well tolerated. Both GLP-1 and GIP mimetics have shown neuroprotective properties in animal models of Parkinson's and Alzheimer's disease. The aim of this study is to evaluate the potential neuroprotective effects of liraglutide, a GLP-1 analog, and twincretin, a dual GLP-1R/GIPR agonist, in a murine mTBI model. First, we subjected mice to mTBI using a weight-drop device and, thereafter, administered liraglutide or twincretin as a 7-day regimen of subcutaneous (s.c.) injections. We then investigated the effects of these drugs on mTBI-induced cognitive impairments, neurodegeneration, and neuroinflammation. Finally, we assessed their effects on neuroprotective proteins expression that are downstream to GLP-1R/GIPR activation; specifically, PI3K and PKA phosphorylation. Both drugs ameliorated mTBI-induced cognitive impairments evaluated by the novel object recognition (NOR) and the Y-maze paradigms in which neither anxiety nor locomotor activity were confounds, as the latter were unaffected by either mTBI or drugs. Additionally, both drugs significantly mitigated mTBI-induced neurodegeneration and neuroinflammation, as quantified by immunohistochemical staining with Fluoro-Jade/anti-NeuN and anti-Iba-1 antibodies, respectively. mTBI challenge significantly decreased PKA phosphorylation levels in ipsilateral cortex, which was mitigated by both drugs. However, PI3K phosphorylation was not affected by mTBI. These findings offer a new potential therapeutic approach to treat mTBI, and support further investigation of the neuroprotective effects and mechanism of action of incretin-based therapies for neurological disorders.

Interaction between methylphenidate, methadone and different antidepressant drugs on antinociception in mice, and possible clinical implications.

OBJECTIVES: Methylphenidate (MPH), a psychostimulant used for treatment of attention deficit hyperactivity disorder (ADHD), is widely used by patients on antidepressants and methadone maintenance treatment (MMT). Preclinical studies showed MPH to exert analgesic effects when given alone or with morphine. METHODS: Using the hotplate assay on mice, we studied the interaction of acute doses of MPH with sub-threshold doses of methadone and different antidepressant medications and the interaction of increasing doses of MPH with chronic methadone. RESULTS: Adding a sub-threshold dose of venlafaxine, desipramine or clomipramine to MPH produced significant augmentation of MPH antinociception with each medication (P < 0.05). No such interactions were found between escitalopram and acute methadone. However, addition of increasing doses of MPH to chronic methadone given for 2 weeks using ALZET osmotic mini pumps induced augmentation of the antinociceptive effect of chronic methadone exclusively at high dose of MPH (7.5 mg/kg). CONCLUSIONS: These findings may implicate the need of an excessive attention to the administration of MPH to MMT patients. The no interaction found between MPH and escitalopram may hint to the possibly safe co-administration of MPH and selective serotonin reuptake inhibitors (SSRIs) to depressed ADHD patients. Further studies are needed in order to validate these possible clinical implications.

The Invisibility of Mild Traumatic Brain Injury: Impaired Cognitive Performance as a Silent Symptom.

The present study was designed to tackle two notorious features of mild traumatic brain injury (mTBI)-heterogeneity and invisibility-by characterizing the full scope of mTBI symptoms. Mice were exposed to brain injuries of different intensities utilizing a weight-drop model (10, 30, 50, and 70 g) and subsequently subjected to a comprehensive battery of behavioral tests at different time points and immunohistochemical examination of cortical slices. Whereas the physiological, neurological, emotional, and motor function of mTBI mice (i.e., their well-being) remained largely intact, cognitive deficits were identified by the y-maze and novel object recognition. Results from these two cognitive tests were combined and a dose-response relationship was established between injury intensity and cognitive impairment, ranging from an 85% decline after a 70-g impact (p < 0.001) to a 20% decline after a 10-g impact (essentially no effect). In addition, higher intensities of injury were accompanied by decreased expression of axonal and synaptic markers. Thus, our mTBI mice showed a clear discrepancy between performance (poor cognitive function) and appearance (healthy demeanor). This is of major concern given that diagnosis of mTBI is established on the presence of clinical symptoms and emphasizes the need for an alternative diagnostic modality.

Novel GLP-1R/GIPR co-agonist "twincretin" is neuroprotective in cell and rodent models of mild traumatic brain injury.

Several single incretin receptor agonists that are approved for the treatment of type 2 diabetes mellitus (T2DM) have been shown to be neuroprotective in cell and animal models of neurodegeneration. Recently, a synthetic dual incretin receptor agonist, nicknamed "twincretin," was shown to improve upon the metabolic benefits of single receptor agonists in mouse and monkey models of T2DM. In the current study, the neuroprotective effects of twincretin are probed in cell and mouse models of mild traumatic brain injury (mTBI), a prevalent cause of neurodegeneration in toddlers, teenagers and the elderly. Twincretin is herein shown to have activity at two different receptors, dose-dependently increase levels of intermediates in the neurotrophic CREB pathway and enhance viability of human neuroblastoma cells exposed to toxic concentrations of glutamate and hydrogen peroxide, insults mimicking the inflammatory conditions in the brain post-mTBI. Additionally, twincretin is shown to improve upon the neurotrophic effects of single incretin receptor agonists in these same cells. Finally, a clinically translatable dose of twincretin, when administered post-mTBI, is shown to fully restore the visual and spatial memory deficits induced by mTBI, as evaluated in a mouse model of weight drop close head injury. These results establish twincretin as a novel neuroprotective agent and suggest that it may improve upon the effects of the single incretin receptor agonists via dual agonism.