Fragile X Syndrome (FXS) is the most common inherited cause of intellectual disability and is the leading known single-gene cause of autism spectrum disorder. FXS patients display varied behavioural deficits that include mild to severe cognitive impairments in addition to mood disorders. Currently, there is no cure for this condition, however, there is an emerging focus on therapies that inhibit mTOR-dependent protein synthesis due to the clinical effectiveness of metformin for alleviating some behavioural symptoms in FXS. Adiponectin (APN) is a neurohormone that is released by adipocytes and provides an alternative means to inhibit mTOR activation in the brain. In these studies, we show that Fmr1 KO mice, like FXS patients, show reduced levels of circulating APN, and that both LTP and LTD in the DG (dentate gyrus) are impaired. Brief (20 min) incubation of hippocampal slices in APN (50 nM) was able to rescue both LTP and LTD in the DG and increased both the surface expression and phosphorylation of GluA1 receptors. These results provide evidence for reduced adiponectin levels in FXS playing a role in decreasing bidirectional synaptic plasticity and show that therapies that enhance adiponectin levels may have therapeutic potential for this and related conditions.

Slices were perfused with ACSF (30 °C) at a rate of approximately 2 ml/min and field excitatory postsynaptic potentials (fEPSPs) recorded using an Axon MultiClamp 700B amplifier connected to a PC running Clampex 10.2 software (Molecular Devices, CA, USA). An upright Olympus BX50WI microscope was used to assist with visually placing stimulating and recording electrodes in the middle of the molecular lay of the DG as previously described. fEPSPs were elicited by delivering a 120 μs (10–40 μA) current pulse every 15s to the medial perforant path (MPP) via a concentric bipolar stimulating electrode (FHC, Bowdoin, ME). A borosilicate glass recording electrode (1 MΩ) was filled with ACSF and visually placed about 200 μm from the stimulating electrode. A stable baseline was recorded for at least 15 min using a stimulation magnitude that elicit a response of 50% of the maximum amplitude obtained for synaptic potentiation, and 70% for synaptic depression experiments. Following this, a paired-pulse protocol and an input-output (I/O) protocols were applied to evaluate possible changes in synaptic transmission induced by the experimental variables investigated. Paired-pulse experiments consisted in the application of 6 sets of 2 pulses each with a 50 ms interpulse interval, and a ratio was calculated by dividing the slope of the second fEPSP by the first.

In order to induce LTP induction in the DG, slices were exposed to the GABA_A receptor antagonist bicuculline methiodide during baseline recordings (5 μM washed for 20 min; Sigma-Aldrich, ON, Canada). This protocol allows for a significant reduction in tonic inhibition and isolation of the excitatory component of synaptic transmission in this hippocampal subregion. In addition, to assess the impact of long-term exposure to APN (50 nM; >90 min incubation), slices were constantly exposed to ACSF supplemented with this hormone during both recovery time and recordings. Short-time exposure (50 nM; 20 min incubation) was evaluated by exposing slices to APN diluted in ACSF for 20 min during baseline recordings (in addition to bicuculline methiodide for LTP experiments). LTP was induced by applying a high-frequency stimulus (HFS; 4 trains of 50 pulses at 100 Hz, 30 s apart), and LTD by prolonged low-frequency stimulation (LFS; 900 pulses at 1 Hz, 1 s apart). Responses were recorded for 60 min after the conditioning stimulus, and fEPSPs slopes were calculated using Clampfit 10 (Axon Instruments). Two parameters were then quantified for statistical comparisons: LTP was calculated ~60 min following induction by averaging 20 successive recordings; PTP (post-tetanic potentiation) was calculated by averaging the first 4 responses following induction. (This measure includes a mixture of a rapid NMDAR-independent potentiation and NMDAR-dependent short-term potentiation; STP). Experiments were conducted using hippocampal slices that exhibited response amplitudes exceeding 1 mV and a favorable fEPSP to fiber volley ratio of 2:1 or higher, which was considered qualitative indicators of robust tissue responses.