The effects of sigma (σ1) receptor-selective ligands on muscarinic receptor antagonist-induced cognitive deficits in mice

Abstract

Background and purpose: Cognitive deficits in patients with Alzheimer's disease, Parkinson's disease, traumatic brain injury and stroke often involve alterations in cholinergic signalling. Currently available therapeutic drugs provide only symptomatic relief. Therefore, novel therapeutic strategies are needed to retard and/or arrest the progressive loss of memory.

Experimental approach: Scopolamine-induced memory impairment provides a rapid and reversible phenotypic screening paradigm for cognition enhancement drug discovery. Male C57BL/6J mice given scopolamine (1 mg·kg(-1) ) were used to evaluate the ability of LS-1-137, a novel sigma (σ1) receptor-selective agonist, to improve the cognitive deficits associated with muscarinic antagonist administration.

Key results: LS-1-137 is a high-affinity (Ki = 3.2 nM) σ1 receptor agonist that is 80-fold selective for σ1, compared with σ2 receptors. LS-1-137 binds with low affinity at D2-like (D2, D3 and D4) dopamine and muscarinic receptors. LS-1-137 was found to partially reverse the learning deficits associated with scopolamine administration using a water maze test and an active avoidance task. LS-1-137 treatment was also found to trigger the release of brain-derived neurotrophic factor from rat astrocytes.

Conclusions and implications: The σ1 receptor-selective compound LS-1-137 may represent a novel candidate cognitive enhancer for the treatment of muscarinic receptor-dependent cognitive deficits.

Figure 1

LS-1–137 regulation of σ1 receptor-BiP complex formation. The efficacy of LS-1–137 was compared with the known σ1 receptor agonist PRE-084 and antagonist NE-100. Vehicle (control) or test drugs were applied into culture medium for 30 min at 37°C. Cell lysates were collected and used for the detection of σ1 receptors coupled to BiP. Test drugs were applied at 1 or 10 μM final concentration (n ≥ 3). Data is presented as the percent (%) of untreated control cells ± SEM. ***P = 0.004 versus control; ****P < 0.001 versus control; ns P > 0.05 not significant.

Figure 2

Modulation of scopolamine effects on learning by LS-1–137 using the active avoidance task. An active avoidance task was used to assess the effects of scopolamine on short-term learning. All test drugs were administered prior to starting the test. Male C57BL/6J mice were pretreated with LS-1–137 (LS) or vehicle (Veh) at 15 min prior to scopolamine (Scop) administration. The test was started 30 min after the scopolamine injection. The data presented in the figure panels represents the number of trials for an animal to meet the criteria. Data are reported as the means ± SEM. for n ≥ 6 animals per group. *P < 0.05 vs. scopolamine and #P < 0.001 vs. vehicle group). Panel A (left): comparison of the effects of vehicle versus scopolamine (1 mg·kg⁻¹) versus scopolamine plus LS-1–137 (3 mg·kg⁻¹) versus vehicle plus LS-1–137. Panel B (middle): comparison of the effects of vehicle versus scopolamine (1 mg·kg⁻¹) versus scopolamine in the presence of PRE-084, PPCC or NE-100. Panel C (right): ability of NE-100 to attenuate the inhibition of the effect of scopolamine by LS-1–137. *P < 0.05, **P < 0.008.

Figure 3

Effect of scopolamine (Scop) and LS-1–137 on latency, path length and speed in Morris water maze. Male C57BL/6J mice were pretreated with LS-1–137 (LS; 3 mg·kg⁻¹) 15 min prior to Scop administration (1 mg·kg⁻¹). Testing was started 30 min after Scop administration. Top panel: representative swim paths of animals in different groups (left to right: vehicle control vs. Scop vs. Scop plus LS-1–137 (LS) vs. LS-1–137) is shown. Bottom panel: data are presented as the mean ± SEM for n ≥ 8 animals per group for the daily effects of scopolamine, vehicle control, LS-1–137 in the presence of scopolamine or LS-1–137. Bottom left panel: time taken to reach the platform (latency in seconds). Bottom middle panel: average distance travelled to the platform (path length in cm). Bottom right panel: swim speed in cm·s⁻¹. *Compared with Scop-treated group, #compared with vehicle control group. P ≤ 0.05 is considered significant.

Figure 4

Comparison of the binding of scopolamine and LS-1–137 to muscarinic receptors using ³H-QNB and rat cortical brain tissue. Competitive radioligand binding studies were conducted using murine brain cortex tissue and ³H-QNB to label muscarinic receptors. Non-specific binding was defined using 1 μM atropine. This figure shows a composite competition curve for the specific binding of ³H-QNB as a function of inhibitor concentration, where each point is the mean inhibition from n = 4 for scopolamine and n = 3 for LS-1–137. IC50 values for each independent experiment were converted to K_i values using the Cheng and Prusoff equation (1973). The mean K_i values ± SEM for ³H-QNB binding sites were found to be 1447 ± 894 nM (n = 3) for LS-1–137 and 0.61 ± 0.16 nM (n = 4) for scopolamine.

Figure 5

Effect of LS-1–137 on rat recombinant GluN1GluN2A NMDA receptors transiently expressed in HEK 293 cells. (A, top) Representative traces of whole-cell NMDA (20 μM)-activated currents recorded from same cell is shown. LS-1–137 (1 or 10 μM) was co-applied with NMDA to the cell for 10 s. (Bottom) Summary data of the effect of LS-1–137 on NMDA response are shown. (B, top) Representative recording of NMDA-activated currents recorded before, during the 3 min perfusion with 10 μM LS-1–137 and the after washout is shown. (Bottom) Summary data of effect of LS-1–137 treatment on NMDA response are shown. All current amplitudes are normalized to NMDA response in the absence of LS-1–137 (as 100%). Each data point represents mean ± SEM from at ≥4 cells.

Figure 6

LS-1–137 induced BDNF release from rat astrocytes. LS-1–137 induced BDNF release in a concentration-dependent manner from rat primary cortical astrocytes. Cells were treated with LS-1–137 at the indicated concentrations for 18 h. BDNF release into the media was measured by elisa. The data are representative of three independent experiments and expressed as the mean ± SEM BDNF (pg·mL⁻¹). The mean baseline for this assay (vehicle control) was 1.51 ± 0.18 pg·mL⁻¹.