Aberrant hippocampal Ca2+ micro-waves following synapsin-dependent adeno-associated viral expression of Ca2+ indicators.

Genetically encoded calcium indicators (GECIs) such as GCaMP are invaluable tools in neuroscience to monitor neuronal activity using optical imaging. The viral transduction of GECIs is commonly used to target expression to specific brain regions, can be conveniently used with any mouse strain of interest without the need for prior crossing with a GECI mouse line and avoids potential hazards due to the chronic expression of GECIs during development. A key requirement for monitoring neuronal activity with an indicator is that the indicator itself minimally affects activity. Here, using common adeno-associated viral (AAV) transduction procedures, we describe spatially confined aberrant Ca2+ micro-waves slowly travelling through the hippocampus following expression of GCaMP6, GCaMP7 or R-CaMP1.07 driven by the synapsin promoter with AAV-dependent gene transfer, in a titre-dependent fashion. Ca2+ micro-waves developed in hippocampal CA1 and CA3, but not dentate gyrus (DG) nor neocortex, were typically first observed at 4 weeks after viral transduction, and persisted up to at least 8 weeks. The phenomenon was robust, observed across laboratories with various experimenters and setups. Our results indicate that aberrant hippocampal Ca2+ micro-waves depend on the promoter and viral titre of the GECI, density of expression as well as the targeted brain region. We used an alternative viral transduction method of GCaMP which avoids this artifact. The results show that commonly used Ca2+-indicator AAV transduction procedures can produce artefactual Ca2+ responses. Our aim is to raise awareness in the field of these artefactual transduction-induced Ca2+ micro-waves and we provide a potential solution.

Sustained attention performance deficits in the three-choice serial reaction time task in male and female rats after experimental brain trauma.

Impaired attention is central to the cognitive deficits associated with long-term sequelae for many traumatic brain injury (TBI) survivors. Assessing complex sustained attention post-TBI is clinically-relevant and may provide reliable avenues towards developing therapeutic and rehabilitation targets in both males and females. We hypothesized that rats subjected to a moderate TBI will exhibit attentional deficits seen as reduced accuracy and increased distractibility in an operant 3-choice serial reaction time task (3-CSRT), designed as an analogue of the clinical continuous performance test. Upon reaching baseline of 70% accuracy at the 300 ms cue, adult male and female Sprague-Dawley rats were subjected to a controlled cortical impact (2.8 mm deformation at 4 m/s) or sham injury over the right parietal cortex. After two weeks of recovery, they were retested on the 3-CSRT for ten days. Dependent measures include percent accuracy (overall and for each of the three cue ports), percent omissions, as well as latency to instrumental poke and retrieve reward. Results demonstrate that both males and females displayed reduced percent accuracy and increased omissions when re-tested post-TBI on 3-CSRT compared to Sham rats and to their own pre-insult baseline (p's < 0.05). Performance accuracy was impaired consistently throughout the ten days of post-surgery re-testing, suggesting pronounced and long-lasting dysfunction in sustained attention processes. Deficits were specifically more pronounced when the cue was pseudorandomly presented in the left-side cue port (p < 0.05), mirroring clinical hemispatial neglect. These data demonstrate significant and persistent complex attention impairments in both sexes after TBI, rendering identifying efficient therapies for cognitive recovery as pivotal.

Milnacipran Ameliorates Executive Function Impairments following Frontal Lobe Traumatic Brain Injury in Male Rats: A Multimodal Behavioral Assessment.

Traumatic brain injuries (TBIs) affect more than 10 million patients annually worldwide, causing long-term cognitive and psychosocial impairments. Frontal lobe TBIs commonly impair executive function, but laboratory models typically focus primarily on spatial learning and declarative memory. We implemented a multi-modal approach for clinically relevant cognitive-behavioral assessments of frontal lobe function in rats with TBI and assessed treatment benefits of the serotonin-norepinephrine reuptake inhibitor, milnacipran (MLN). Two attentional set-shifting tasks (AST) evaluated cognitive flexibility via the rats' ability to locate food-based rewards by learning, unlearning, and relearning sequential rule sets with shifting salient cues. Adult male rats reached stable pre-injury operant AST (oAST) performance in 3-4 weeks, then were isoflurane-anesthetized, subjected to a unilateral frontal lobe controlled cortical impact (2.4 mm depth, 4 m/sec velocity) or Sham injury, and randomized to treatment conditions. Milnacipran (30 mg/kg/day) or vehicle (VEH; 10% ethanol in saline) was administered intraperitoneally via implanted osmotic minipumps (continuous infusions post-surgery, 60 µL/h). Rats had a 10-day recovery post-TBI/Sham before performing light/location-based oAST for 10 days and, subsequently, odor/media-based digging AST (dAST) on the last test day (26-27 days post-injury) before sacrifice. Both AST tests revealed significant deficits in TBI+VEH rats, seen as elevated total trials and errors (p < 0.05), which generally normalized in MLN-treated rats (p < 0.05). This first simultaneous dual AST assessment demonstrates oAST and dAST are sufficiently sensitive and robust to detect subtle attentional and cognitive flexibility executive impairments after frontal lobe TBI in rats. Chronic MLN administration shows promise for attenuation of post-TBI executive function deficits, thus meriting further investigation.

Chronic treatment with galantamine rescues reversal learning in an attentional set-shifting test after experimental brain trauma.

Approximately 10 million new cases of traumatic brain injury (TBI) are reported each year worldwide with many of these injuries resulting in higher order cognitive impairments. Galantamine (GAL), an acetylcholine esterase inhibitor (AChEI) and positive allosteric modulator of nicotinic acetylcholine receptors (nAChRs), has been reported to ameliorate cognitive deficits after clinical TBI. Previously, we demonstrated that controlled cortical impact (CCI) injury to rats resulted in significant executive function impairments as measured by the attentional set-shifting test (AST), a complex cognitive task analogous to the Wisconsin Card Sorting Test (WCST). We hypothesized that chronic administration of GAL would normalize performance on the AST post-TBI. Isoflurane-anesthetized adult male rats were subjected to moderate CCI (2.8 mm tissue deformation at 4 m/s) or sham injury. Rats were then randomized into one of three treatment groups (i.e., 1 mg/kg GAL, 2 mg/kg GAL, or 1 mL/kg saline vehicle; VEH) or their respective sham controls. GAL or VEH was administered intraperitoneally daily commencing 24 hours post-surgery and until AST testing at 4 weeks post-injury. The AST data revealed significant impairments in the first reversal stage after TBI, seen as increased trials to reach criterion and elevated total errors (p < 0.05). These behavioral flexibility deficits were equally normalized by the administration of both doses of GAL (p < 0.05). Additionally, the higher dose of GAL (2 mg/kg) also significantly reduced cortical lesion volume compared to TBI + VEH controls (p < 0.05). In summary, daily GAL administration provides an efficacious treatment for cognitive deficits and histological recovery after experimental brain trauma. Clinically, these findings are promising considering robust results were attained using a pharmacotherapy already used in the clinic to treat mild dementia.

Spontaneous recovery of traumatic brain injury-induced functional deficits is not hindered by daily administration of lorazepam.

Agitation and aggression are common sequelae of traumatic brain injury (TBI) and pose a challenge to physicians and other health providers during acute patient care and subsequent neurorehabilitation. Antipsychotic drugs (APDs) are routinely administered to manage TBI patients displaying such maladaptive behaviors despite several clinical and preclinical studies demonstrating that they hinder recovery. A potentially viable alternative to APDs may be the benzodiazepines, which have differing mechanisms of action. Hence, the aim of the study was to test the hypothesis that lorazepam (LOR) would not impede recovery after TBI. Anesthetized adult male rats received a cortical impact or sham injury and then were intraperitoneally administered LOR (0.1mg/kg, 1.0mg/kg, or 2.0mg/kg) or vehicle (VEH; 1mL/kg) commencing 24-h after surgery and once daily for 19days. Motor and cognitive outcomes were assessed on post-operative days 1-5 and 14-19, respectively. No differences were revealed among the four sham control groups and thus they were pooled into one inclusive SHAM group. The SHAMs performed better than all TBI groups on all assessments (p<0.05). Regarding TBI, the 2.0mg/kg LOR group performed better than the VEH and 0.1mg/kg or 1.0mg/kg LOR groups on every task (p<0.05); no differences were observed among the latter three groups on any endpoint (p>0.05). Overall, these preclinical behavioral data support the hypothesis and reveal a therapeutic benefit with the higher dose of LOR. The findings suggest that LOR may be an alternative, to APDs, for controlling agitation without compromising spontaneous recovery and perhaps could afford a dual benefit by also promoting therapeutic efficacy.

Abbreviated environmental enrichment confers neurobehavioral, cognitive, and histological benefits in brain-injured female rats.

Environmental enrichment (EE) promotes behavioral recovery after experimental traumatic brain injury (TBI). However, the chronic rehabilitation provided in the laboratory is not analogous to the clinic where physiotherapy is typically limited. Moreover, females make up approximately 40% of the clinical TBI population, yet they are seldom studied in brain trauma. Hence, the goal of this study was to test the hypothesis that abbreviated EE would confer neurobehavioral, cognitive, and histological benefits in brain injured female rats. Anesthetized rats received a cortical impact of moderate-to-severe injury (2.8mm tissue deformation at 4m/s) or sham surgery and then were randomly assigned to groups receiving standard (STD) housing or 4h, 6h, or 24h of EE daily. Motor function (beam-balance/walk and rotarod) was assessed on post-operative days 1-5 and every other day from 1 to 19, respectively. Spatial learning/memory (Morris water maze) was evaluated on days 14-19, and cortical lesion volume was quantified on day 21. No statistical differences were appreciated among the sham controls in any assessment and thus the data were pooled. All EE conditions improved motor function and memory retention, but only 6h and 24h enhanced spatial learning relative to STD (p<0.05). Moreover, EE, regardless of duration reduced cortical lesion volume (p<0.05). These data confirm that abbreviated EE confers robust neurobehavioral, cognitive, and histological benefits in TBI female rats, which supports the hypothesis and strengthens the utility of EE as a pre-clinical model of neurorehabilitation.