Learning deficits in rats overexpressing the dopamine transporter

Abstract

With its capacity to modulate motor control and motivational as well as cognitive functions dopamine is implicated in numerous neuropsychiatric diseases. The present study investigated whether an imbalance in dopamine homeostasis as evident in the dopamine overexpressing rat model (DAT-tg), results in learning and memory deficits associated with changes in adult hippocampal neurogenesis. Adult DAT-tg and control rats were subjected to the Morris water maze, the radial arm maze and a discrimination reversal paradigm and newly generated neurons in hippocampal circuitry were investigated post mortem. DAT-tg rats were found to exhibit a striking inability to acquire information and deploy spatial search strategies. At the same time, reduced integration of adult-born neurons in hippocampal circuitry was observed, which together with changes in striatal dopamine signalling might explain behavioural deficits.

Figure 1

Experimental design. Labelling of newly generated cells for analysis of hippocampal neurogenesis was achieved by injecting proliferation marker BrdU into rats, three times with a six hours interval at a dose of 50 mg/kg. Following BrdU injections, animals performed a series of tasks investigating aspects of learning and memory and sensorimotor function. Behavioural experiments were performed in 3 batches of animals (A: wt n = 4, het n = 8; B: wt n = 2, het n = 4;C: wt n = 5, het n = 4). Test order Batch A: RAM, DR, SCT, OF/OP/NOR, MWM; Batch B: RAM, OF/OP/NOR, MWM, SCT, DR; Batch C: MWM, RAM, OF/OP/NOR, SCT, DR. BrdU = 5-Bromo-2′Deoxyuridine, MWM = Morris water maze, DR = Discrimination Learning, RAM = Radial arm maze, OF = open –field, OP = open platform, NOR = novel object recognition, SCT = sucrose consumption test, PND = postnatal day, IHC = immunohistochemistry.

Figure 2

Behavioural assessment of learning and memory. (d) Schematics of MWM set up. In the Acquisition phase (day1–4) DAT-tg rats do not learn to locate the hidden platform (a), thereby showing longer path length compared to controls (b) but intact motor function (velocity (c)). (e) The probabilistic occupancy plots represent sum data over trials and rats within respective groups and illustrate the rapid development of a place-specific preference for the platform position for control animals but not DAT-tg animals. (e–g) In accordance to the impairment in learning during acquisition DAT-tg animals do not prefer the former goal quadrant after platform removal as found for controls. (e) During probe trial DAT-tg rats continuously show thigmotactic swimming around the wall of the pool. DAT-tg: n = 16; wt: n = 11 (h,i) Performance during cued platform trials indicates intact sensorimotor function when platform is visible. DAT-tg: n = 10; wt: n = 8 (j) During the discrimination learning paradigm DAT-tg rats exhibit impairments in initial acquisition. DAT-tg: n = 9; wt: n = 11 (k) During RAM DAT-tg rats exhibit significantly longer trial durations due to lack in exploratory behaviour. DAT-tg: n = 16; wt: n = 11 Dashed line (a,f) indicates chance level. Error bars indicate the standard error of the mean, significance level p < 0.05.

Figure 3

Exploratory behaviour and general locomotor activity. Results from the (a–c) Open-field (OF) (d–f) Open-platform (OP) (g–i) novel object recognition (NOR) and (j) Sucrose consumption test (SCT) are presented. (i) Discrimination index can vary between +1 and −1, where a positive score indicates more time spent with the novel object for controls, and a negative score for DAT-tg rats indicates more time spent with the familiar object. The dashed line indicates a null preference of novel-object investigation. (a–f,j) DAT-tg: n = 16; wt: n = 11 (g–i) DAT-tg: n = 7; wt: n = 11 Error bars indicate the standard error of the mean, significance level p < 0.05.

Figure 4

Histochemical analysis of hippocampal neurogenesis. Ten weeks after injection and behavioural assessment proliferation (Ki67) survival of newly generated cells (BrdU) and proportion of generated neurons (BrdU/NeuN) was quantified. The number of (a) Ki67-positive cells for DAT-tg: n = 9; wt: n = 7 and (b) BrdU-positive cells. DAT-tg: n = 13; wt: n = 7 did not differ between genotypes. However in DAT-tg rats lower numbers of BrdU/NeuN double-labelled cells could be detected. DAT-tg: n = 13; wt: n = 7. Error bars indicate the standard error of the mean, significance level p < 0.05. (d–g) Representative images for Ki67 DAB staining (d,e) and BrdU/NeuN immunofluorescent staining (f,g) are shown. Ki67 bright field, NeuN, red; BrdU, green; Scale bar: (d,e) 150 µm, (f,g) 100 µm, (insets) 15 µm.