Neuroprotective Effects of a Serotonin Receptor Peptide Following Sham vs. Mild Traumatic Brain Injury in the Zucker Rat

Abstract

Aims: Accelerated cognitive decline frequently complicates traumatic brain injury. Obesity and type 2 diabetes mellitus drive peripheral inflammation which may accelerate traumatic brain injury-associated neurodegeneration. The Zucker rat harbors G-protein coupled receptor agonist IgG autoantibodies and in vitro neurotoxicity caused by these autoantibodies was prevented by a novel synthetic fragment of the serotonin 2A receptor. The aim of the present study was to test whether genetic obesity manifested in Zucker diabetic fatty rat is associated with greater spatial memory impairment before and after mild traumatic brain injury compared to Zucker lean rats. Furthermore, we investigated whether these neurodegenerative complications can be lessened by administration of a novel putative neuroprotective peptide comprised of a fragment of the second extracellular loop of the serotonin 2A receptor.

Methods: Age-matched lean and fatty diabetic Zucker rats were tested in the Morris water maze (spatial memory) prior to receiving a sham-injury or lateral fluid percussion (LFP) mild traumatic brain injury. Behavioral testing was repeated at 1-week, 1-month, and 3-month intervals following injury. A synthetic peptide consisting of a portion of the 5-hydroxytryptamine (serotonin) 2A receptor (2 mg/kg) (vehicle, or an inactive scrambled version of the peptide (2 mg/kg)) was administered via intraperitoneal route every other day for 7 days after sham or LFP injury to lean rats or 7 days before and after sham or LFP injury to fatty rats.

Results: Mild traumatic brain injury impaired recall of spatial memory in fatty and lean rats. Zucker fatty rats subjected to sham-injury or mild TBI experienced a significantly greater longitudinal decline in recall of spatial memory compared to lean Zucker rats. A synthetic peptide fragment of the 5-hydroxytryptamine 2A receptor significantly enhanced acquisition of spatial learning and it appeared to strengthen recall of spatial learning (one-week) after sham injury in Zucker rats.

Conclusions: These data suggest that the Zucker diabetic fatty rat is a suitable animal model to investigate the role of metabolic factor(s) in accelerated cognitive decline. A novel synthetic peptide comprised of a fragment of the second extracellular loop of the human serotonin 2A receptor appeared to have neuroprotective effects on both acquisition and recall of spatial memory in subsets of Zucker rats, with relatively greater benefit in sham-injured, lean Zucker rats.

Figure 1:

Timeline of treatments, exposures and behavioral assessments in Zucker rats.

Figure 2:

Delayed match to position in the Morris water maze: distance (meters) as a function of time before and after injury in Zucker lean and fatty rats. Results are mean ± SEM. Distance was determined by Any Maze software as described in Methods. Fatty: sham (N=6), tbi (N=6) Lean: Sham (N=9), tbi (N=8)

Figure 3:

Delayed match to position in the Morris water maze: path efficiency as a function of time before and after injury in Zucker lean and fatty rats. Results are mean ± SEM. Path efficiency was determined by Any Maze software as described in Methods. Fatty (N=12); Lean (N=17) rats.

Figure 4:

Delayed match to position in the Morris water maze: Path efficiency difference at 1-week post injury in Zucker fatty and lean rats. Path efficiency difference was calculated as described in Methods and is a measure of the strength of recall of spatial learning. Results are mean ± SEM. Sham (N=9); TBI (N=11). Sham, saline (N=5), Sham, P4 (N=4), TBI, saline (N=4), TBI, P4 (N=7). P4= SN..8 peptide.

Figure 5:

Delayed match to position in the Morris water maze: duration in the choice phase CP (recall of spatial learning) as a function of time before and after injury in Zucker lean (N=12) and fatty rats (N=22). Results are mean ± SEM. Duration (seconds) was determined by Any Maze software as described in Methods.

Figure 6:

Delayed match to position in the Morris water maze: duration in the sample phase SP (acquisition of spatial learning) as a function of time, A) and injury or B) peptide (P4= SN..8, scrambled = LD..8) treatment group or C) strain before and after injury in Zucker lean (N=12) and fatty rats (N=22). Results are mean ± SEM. Duration was determined by Any Maze software as described in Methods.

Figure 7:

Dose-dependent inhibition of N2A neurite retraction from 100 nanomolar concentration of Zucker heterozygous lean by SN..8. Neurite retraction assay were performed as described in Methods. Results are mean ± SEM.

Figure 8:

Effect of targeted ‘C to A’ or ‘D to A’ amino acid substitutions in ‘wild-type’ SCLLADDN peptide on the resulting mutant peptides’ (SALLADDN) or (SCLLADAN) ability to prevent acute N2A neurite retraction in the presence of 50–100 nanomolar concentration of Zucker heterozygous lean rat IgG. Results are mean ± SEM of three experiments. Similar results were obtained in experiments using the IgG fraction of plasma from four different middle-aged human TBI patients. Neurite retraction assay in mouse N2A neuroblastoma cells was carried out as described in Methods.