Sex Differences in the Cholinergic Basal Forebrain in the Ts65Dn Mouse Model of Down Syndrome and Alzheimer's Disease

Abstract

In the Down syndrome (DS) population, there is an early incidence of dementia and neuropathology similar to that seen in sporadic Alzheimer’s disease (AD), including dysfunction of the basal forebrain cholinergic neuron (BFCN) system. Using Ts65Dn mice, a model of DS and AD, we examined differences in the BFCN system between male and female segmentally trisomic (Ts65Dn) and disomic (2N) mice at ages 5–8 months. Quantitative stereology was applied to BFCN subfields immunolabeled for choline acetyltransferase (ChAT) within the medial septum/vertical limb of the diagonal band (MS/VDB), horizontal limb of the diagonal band (HDB) and nucleus basalis of Meynert/ substantia innominata (NBM/SI). We found no sex differences in neuron number or subregion area measurement in the MS/VDB or HDB. However, 2N and Ts65Dn females showed an average 34% decrease in BFCN number and an average 20% smaller NBM/SI region area compared with genotype-matched males. Further, relative to genotype-matched males, female mice had smaller BFCNs in all subregions. These findings demonstrate that differences between the sexes in BFCNs of young adult Ts65Dn and 2N mice are region and genotype specific. In addition, changes in post-processing tissue thickness suggest altered parenchymal characteristics between male and female Ts65Dn mice.

Maternal choline supplementation improves spatial mapping and increases basal forebrain cholinergic neuron number and size in aged Ts65Dn mice

“…Breeder pairs (Ts65Dn female and C57Bl/6J Eicher × C3H/HeSnJ F1 male mice) were purchased from Jackson Laboratories (Bar Harbor, ME), mated at Cornell University, (Ithaca, NY), and randomly assigned to either a choline-controlled, standard rodent chow diet (AIN-76A with 1.1 g/kg choline chloride; Dyets Inc., Bethlehem, PA) or a rodent chow diet with choline supplementation (AIN-76A with 5.0 g/kg choline chloride; Dyets Inc) as reported previously (Kelley et al, 2014a, 2014b; Velazquez et al, 2013). The two levels of maternal choline intake selected for these studies with the Ts65Dn model were based on numerous prior studies demonstrating lasting cognitive benefits of increased maternal choline intake in normal rodents (Meck and Williams, 1999; 2003; Meck et al, 2007).…”

“…Brains were extracted from the calvaria, postfixed for 24 h in the same fixative, and placed in a 30% sucrose PB solution at 4 °C until sectioning. Each brain was cut in the coronal plane at 40 μm thickness, on a sliding freezing microtome into six adjacent series and stored at 4 °C in a cryoprotectant solution (30 % ethylene glycol, 30 % glycerol, in 0.1 M PB) prior to immunohistochemical staining (Kelley et al, 2014a, 2014b; Velazquez et al, 2013). …”

“…Immunohistochemistry was performed as previously described (Kelley et al, 2014a, 2014b). Tissue was immunostained using the following primary antibodies: a goat polyclonal antibody against choline acetyltransferase (ChAT, 1:1000; Millipore, Billerica, MA), a rabbit polyclonal antibody against the pan-neurotrophin receptor p75 NTR (1:2000; Millipore), and a rabbit polyclonal antibody against the cognate nerve growth factor (NGF) receptor tyrosine receptor kinase A (TrkA, 1:10,000, gift from Dr. L. Reichardt, UCLA).…”

“…Immunolabeling was visualized using an acetate-imidazole buffer containing 0.05 % 3,3′-diaminobenzidine tetrahy-drochloride (DAB; Sigma-Aldrich, St. Louis, MO) and 0.0015 % freshly prepared H 2 O 2 . Sections were washed in acetate-imidazole buffer to terminate the immunochemical reaction, mounted onto chrome-alum-subbed slides, air dried for 24 h, dehydrated through a series of graded alcohols (70%, 95%, and 100 %), cleared in xylenes, and cover-slipped with distyrene/dibutylphthalate (plasticizer)/xylene (DPX) mounting medium (Kelley et al, 2014a, 2014b; Velazquez et al, 2013). …”

Effects of Maternal Choline Supplementation on the Septohippocampal Cholinergic System in the Ts65Dn Mouse Model of Down Syndrome

“…Mice were deeply anesthetized with ketamine (85 mg/kg)/xylazine (13 mg/kg) via intraperitoneal injection, and perfused transcardially with 0.9 % saline (50 ml), followed by 4% paraformaldehyde (50 ml) in phosphate buffer (PB; 0.1M; pH = 7.4) as previously described [67,68]. Ages at sacrifice are listed in Table 1.…”