Functional perturbation of forebrain principal neurons reveals differential effects in novel and well-learned tasks

Abstract

Neural circuits in mammalian brains consist of large numbers of different cell types having different functional properties. To better understand the separate roles of individual neuron types in specific aspects of spatial learning and memory, we perturbed the function of principal neurons in vivo during maze performance or in hippocampal slices during recording of evoked excitatory synaptic potentials. Transgenic mice expressing the Drosophila allatostatin receptor (AlstR) in cortical and hippocampal pyramidal cells were tested on an elevated plus maze, in a Y-maze, and in the Morris water maze. Relative to a control cohort, AlstR-positive mice treated with allatostatin exhibited no difference in open arm dwell time on the elevated plus maze or total number of arm entries in a Y-maze, but displayed reduced spontaneous alternation. When animals received massed or spaced training trials in the Morris water maze, and the peptide was delivered prior to an immediate probe, no effects on performance were observed. When the peptide was delivered during a probe trial performed 24h after seven days of spaced training, allatostatin delivery to AlstR positive mice enhanced direct navigation to the escape platform. Combined, these results suggest that cortical and hippocampal pyramidal neurons are required during spatial decision-making in a novel environment and compete with other neural systems after extended training in a long-term reference memory task. In hippocampal slices collected from AlstR positive animals, allatostatin delivery produced frequency dependent alterations in the Schaffer collateral fiber volley (attenuated accommodation at 100Hz) and excitatory postsynaptic potential (attenuated facilitation at 5Hz). Combined, the neural and behavioral discoveries support the involvement of short-term plasticity of Schaffer collateral axons and synapses during exploration of a novel environment and during initial orientation to a goal in a well-learned setting.

Keywords: Allatostatin receptor; GIRK; Hippocampus; Pyramidal neuron; Spatial learning and memory.

Fig. 1

Images of colorimetric ISH for tTA and AlstR. A) Complete sagittal images and selected regions from +/+ AlstR mouse brain sections reacted with a tTA probe. B) Complete sagittal images and selected regions from +/+ AlstR mouse brain sections reacted with an AlstR probe. Numbers in top panels indicate distance from midline in mm. Hipp, hippocampus; DMS, dorsomedial striatum; PFC prefrontal cortex; Amyg, amygdala. All images were collected at 10X magnification and tiled (Neurolucida, MBF Bioscience) where necessary.

Fig. 2

Open arm dwell time in the EPM was not affected but spontaneous alternation in a symmetrical Y maze was reduced during silencing of forebrain pyramidal neurons. A) ATD (+/+ receiving allatostatin) and CON animals (all control groups collapsed) spent similar amounts of time in the open arms of the EPM (CON, n = 56; ATD, n = 22). B) Total number of arm entries did not differ across experimental groups (CON, n = 17; ATD, n = 6). C) Alternation rate in the Y maze was reduced in ATD animals compared to CON animals. ^**p < 0.005.

Fig. 3

Silencing of forebrain pyramidal neurons did not affect spatial learning during massed training. A) Escape latencies during training did not differ between CON and ATD animals (CON, n = 40; ATD, n = 14). B) Number of platform crossings during the IMM probe did not differ between CON and ATD animals; C) Plot of quadrant dwell times during the IMM probe shows that neither CON nor ATD animals spent more time in any given quadrant. G = goal, O = opposite, A = adjacent. D) Plot of quadrant biases (percentage of animals spending most of their time in a given quadrant) during the IMM probe shows that neither CON nor ATD animals exhibited a quadrant bias. E) DtP measured across 10 or 30 s of the IMM probe did not differ between CON and ATD animals. The black circle on the Y-axis indicates the platform edge. The open circle marks starting location for all animals (710 mm).

Fig. 4

Silencing of forebrain pyramidal neurons did not affect spatial learning during 8 days of spaced training. A) Escape latencies during training did not differ between CON and ATD animals on Day 5 (CON, n = 38; ATD, n = 14), or Day 8 (CON, n = 40; ATD, n = 12). B) Number of platform crossings during the IMM probe did not differ between CON and ATD animals for Day 5 or Day 8. C) ATD and CON animals in the amount of time animals spent in each quadrant during the IMM probe on Day 5 or Day 8. Both groups spent more time in the goal quadrant than any other quadrant (Chi Square tests); ^*p < 0.05, ^**p < 0.005, ^***p < 0.001. However, only CON animals spent more time in the goal quadrant than any other on Day 8; ^***p < 0.001. D) Both CON and ATD animals showed a bias for the goal quadrant during the IMM probe on Day 5 (CON, n = 38; ATD, n = 14), but only CON showed a preference on Day 8 (CON, n = 37; ATD, n = 11) ^*p < 0.05, ^**p < 0.005. E) DtP measured across 10 s of the IMM probe on Day 5 or Day 8 did not differ between CON and ATD animals. The black circle on the Y-axis indicates the platform edge. The open circle marks starting location for all animals (710 mm). F) There was no difference between CON and ATD animals for DtP on the IMM probe on either Day 5 or Day 8.

Fig. 5

Silencing of forebrain pyramidal neurons improved the initial orientation to the goal location during a 24-HR probe. A) Escape latencies during training did not differ between CON and ATD animals (CON, n = 18; ATD, n = 9). B) Number of platform crossings during the 24-HR probe did not differ between CON and ATD animals during the 24HP probe. C) Both CON and ATD animals showed a preference for the goal quadrant during the 24HP^**p < 0.005 E) DtP measured across 10 s of the 24HP did not differ between CON and ATD animals. The black circle on the Y-axis indicates the platform edge. The open circle marks starting location for all animals (710 mm). F) The first 10 s of the normalized DtP 24HP was reduced for the ATD animals compared to the CON animals [F(1,26) = 6.166, p < 0.05]; * = p < 0.05.

Fig. 6

AlstR activation in hippocampal slices did not alter baseline synaptic transmission but attenuated synaptic facilitation at 5 Hz and AP accommodation at 100 Hz stimulation. One hundred stimulus pulses were applied at 5 (CON, n = 18; ATD, n = 8) or 100 Hz (CON, n = 13; ATD, n = 7). A) Waveforms show all responses across a 100 Hz burst (top, 1 s) or only the first four responses (bottom, 40 ms) for ATD and CON animals. B) Analyses of responses to the first stimulus pulse for all 5 and 100 Hz burst experiments revealed no difference in the mean FP amplitude or mV EPSP slope (CON, n = 18; ATD, n = 12). C) Mean FP amplitude plotted against stimulus number did not differ between groups at 5 Hz stimulation. At 100 Hz, the FP amplitude decreased more during early stimulus pulses (pulse 2–4) for CON slices compared to ATD slices, but decreased more later in the burst (pulses 7–57) for ATD compared to CON slices. * = p < 0.05; *** = p < 0.0001. D) Mean EPSP slope plotted against stimulus number did not differ between groups at 100 Hz stimulation. Synaptic facilitation was reduced in ATD compared to CON animals at 5 Hz. ** = p < 0.001.