Neural Androgen Receptor Deletion Impairs the Temporal Processing of Objects and Hippocampal CA1-Dependent Mechanisms

Abstract

We studied the role of testosterone, mediated by the androgen receptor (AR), in modulating temporal order memory for visual objects. For this purpose, we used male mice lacking AR specifically in the nervous system. Control and mutant males were gonadectomized at adulthood and supplemented with equivalent amounts of testosterone in order to normalize their hormonal levels. We found that neural AR deletion selectively impaired the processing of temporal information for visual objects, without affecting classical object recognition or anxiety-like behavior and circulating corticosterone levels, which remained similar to those in control males. Thus, mutant males were unable to discriminate between the most recently seen object and previously seen objects, whereas their control littermates showed more interest in exploring previously seen objects. Because the hippocampal CA1 area has been associated with temporal memory for visual objects, we investigated whether neural AR deletion altered the functionality of this region. Electrophysiological analysis showed that neural AR deletion affected basal glutamate synaptic transmission and decreased the magnitude of N-methyl-D-aspartate receptor (NMDAR) activation and high-frequency stimulation-induced long-term potentiation. The impairment of NMDAR function was not due to changes in protein levels of receptor. These results provide the first evidence for the modulation of temporal processing of information for visual objects by androgens, via AR activation, possibly through regulation of NMDAR signaling in the CA1 area in male mice.

Fig 1. Western blotting of AR and ERα in the hippocampus of AR^fl/Y and AR^NesCre male mice.

(a-b) Immunoblotting of hippocampal AR (a), ERα (b) and GAPDH proteins (a, b) in control (AR^fl/Y) and mutant (AR^NesCre) males. (c) Quantification of hippocampal ERα protein normalized to GAPDH in 6 males per genotype.

Fig 2. Temporal order memory and object recognition in AR^fl/Y and AR^NesCre male mice.

(a-b) Familiarization and testing phases of the temporal order (a) and novelty detection (b) tasks. (c-d) Discrimination Indexes in the temporal order (c) and novelty detection tasks (d) for males (n = 8–9 males per genotype). ***p < 0.001 versus object A; ^#p < 0.05 versus controls.

Fig 3. Anxiety state level and corticosterone levels in AR^fl/Y and AR^NesCre male mice.

(a-b) Time spent and latency to enter in the open arms of the EPM (a) or the O-maze (b) for controls and mutants (n = 8–11 males per genotype). (f) Corticosterone secretion during the circadian cycle (n = 8–11 males per genotype).

Fig 4. Basal synaptic transmission in the hippocampal CA1 area of AR^fl/Y and AR^NesCre male mice.

(a-b) Input/output (I/O) curves of presynaptic fiber volleys (PFVs) (a) and field excitatory postsynaptic potentials (fEPSPs) (b) induced in control medium by the electrical stimulation of glutamatergic afferents (44 slices from 8 males per genotype). PFV magnitude was significantly smaller in AR^NesCre males only at the highest stimulus intensity (*p < 0.05), whereas fEPSP values were already lower at lower intensities (**p < 0.01). (c) Upper panel, representative paired-pulse facilitation (PPF) of fEPSPs induced in a AR^NesCre male by two successive stimuli (arrows) delivered with a 30 ms inter-stimulus interval. Lower panel, mean PPF magnitude (28–29 slices per genotype).

Fig 5. Synaptic plasticity in the hippocampal CA1 area of AR^fl/Y and AR^NesCre male mice.

(a-b) Comparison of the time course of mean long-term potentiation (LTP) induced by high-frequency stimulation (HFS) (a; 9 slices from 7 animals per genotype) or by theta-burst stimulation (TBS) (b; 12 slices from 7–8 males per genotype). *p < 0.05 versus control. (c) Comparison of the time course of mean long-term depression (LTD) induced by low-frequency stimulation (LFS) in males (10–11 slices from 5–6 animals per genotype).

Fig 6. NMDAR-mediated synaptic activation in the hippocampal CA1 area of AR^fl/Y and AR^NesCre male mice.

(a) Input/Output (I/O) curves of presynaptic fiber volleys (left) and NMDAR-mediated fEPSPs (right) in low-magnesium medium supplemented with 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzoquinoxaline-7-sulfonamide (12–13 slices from 8 males per genotype). PFV magnitude was not affected, whereas fEPSP values were significantly lower at all stimulus intensities in AR^NesCre males (*p < 0.05). (b) Densitometric quantification of NMDAR (GluN1, GluN2A, GluN2B) and non-NMDAR (GluR1, GluR2) proteins normalized to ß-actin in the CA1 area (n = 6–8 males per genotype).