Melanin-concentrating hormone is necessary for olanzapine-inhibited locomotor activity in male mice

Abstract

Olanzapine (OLZ), an atypical antipsychotic, can be effective in treating patients with restricting type anorexia nervosa who exercise excessively. Clinical improvements include weight gain and reduced pathological hyperactivity. However the neuronal populations and mechanisms underlying OLZ actions are not known. We studied the effects of OLZ on hyperactivity using male mice lacking the hypothalamic neuropeptide melanin-concentrating hormone (MCHKO) that are lean and hyperactive. We compared the in vivo effects of systemic or intra-accumbens nucleus (Acb) OLZ administration on locomotor activity in WT and MCHKO littermates. Acute systemic OLZ treatment in WT mice significantly reduced locomotor activity, an effect that is substantially attenuated in MCHKO mice. Furthermore, OLZ infusion directly into the Acb of WT mice reduced locomotor activity, but not in MCHKO mice. To identify contributing neuronal mechanisms, we assessed the effect of OLZ treatment on Acb synaptic transmission ex vivo and in vitro. Intraperitoneal OLZ treatment reduced Acb GABAergic activity in WT but not MCHKO neurons. This effect was also seen in vitro by applying OLZ to acute brain slices. OLZ reduced the frequency and amplitude of GABAergic activity that was more robust in WT than MCHKO Acb. These findings indicate that OLZ reduced Acb GABAergic transmission and that MCH is necessary for the hypolocomotor effects of OLZ.

Keywords: Accumbens nucleus; Anorexia; Antipsychotic; Electrophysiology; Locomotion; MCH.

Figure 1. Acute systemic OLZ treatment suppressed wheel-running activity of WT but not MCHKO mice

A, Mean number of wheel revolutions averaged over 5 days show elevated baseline wheel-running activity of MCHKO mice. B–C, Comparison of cumulative wheel-running activity following intraperitoneal administration of vehicle (0.16% acetic acid) or OLZ (2.5 mg/kg) in WT (B) and MCHKO mice (C) show that OLZ reduced wheel-running activity of WT but not MCHKO mice. D, OLZ-mediated reduction in wheel-running was greater in WT than MCHKO mice. RM-ANOVA for WT vs MCHKO: #, p < 0.05; RM-ANOVA for WT vehicle vs WT OLZ: ^^, p < 0.01; Bonferroni post test: **, p < 0.01.

Figure 2. Bilateral infusion of OLZ into the medial accumbens reduced ambulatory activity of WT but not MCHKO mice

A, Tip location of bilateral cannula placed in the medial Acb for all WT and MCHKO mice included in these experiments. Each cannula pair is color-matched. Ambulatory activity of each WT and MCHKO mouse after intra-Acb OLZ infusion (0.23 µg per side) was compared and normalized to vehicle (0.7% DMSO, 0.01% AcOH). B, Average baseline ambulation count along the x-axis (X-ambulations) over 5 days for MCHKO mice remained elevated following cannulation surgery. C, Comparison of percent decrease in ambulatory activity following intra-Acb vehicle versus OLZ infusion in individual WT and MCHKO mice. Unpaired t test: *, p < 0.05. D, Intra-Acb OLZ infusion reduced ambulatory activity of WT but not MCHKO mice. Inhibitory effects of OLZ in WT Acb were maximal within the first 2 hours but absent across all time points in MCHKO. RM-ANOVA for WT vs MCHKO: #, p < 0.05; Bonferroni post test: *, p < 0.05, **, p < 0.01.

Figure 3. Systemic OLZ treatment reduced GABAergic activity in WT but not MCHKO MSNs ex vivo

MSN recordings from WT and MCHKO Acb were obtained 2–5 hours after mice were injected intraperitoneally with vehicle (0.16% acetic acid; solid bars) or OLZ (2.5 mg/kg; hatched bars). A, Lower MSN sIPSC frequency from OLZ-treated WT but not MCHKO mice. B, MSNs pretreated and recorded in 500 nM TTX showed lower MSN mIPSC frequency from OLZ-treated WT but not MCHKO mice. Unpaired t test: *, p < 0.05, **, p < 0.01.

Figure 4. In vitro OLZ application produced a greater reduction in the frequency of GABAergic activity in WT than MCHKO MSNs

A, Dose-response curve showing the reduction of IPSC frequency by different OLZ concentrations that reached a maximum at 10 µM OLZ. EC₅₀ = 1.6 µM. B, Representative sIPSC sample traces before (control), during (10 µM OLZ), after OLZ washout (wash), and that were abolished by 10 µM bicuculline pretreatment. C, Time course of the change in GABAergic activity by bath application of OLZ showed a robust decrease in sIPSC frequency over time that was greater and longer-lasting in WT than MCHKO MSNs. RM-ANOVA for WT vs MCHKO: #, p < 0.05. D–E, OLZ bath application (dashed lines) to WT and MCHKO MSNs produced a right-shift in the cumulative probability plot of sIPSC (D) and mIPSC (E) interevent intervals compared to their respective controls (solid lines). K-S test for WT and MCHKO control vs OLZ, p < 0.0001. F, Decrease in sIPSC and mIPSC frequency was greater in WT than MCHKO MSNs. Unpaired t test: *, p < 0.05. G–H, Bath application of OLZ (dashed lines) to WT and MCHKO MSNs produced a left-shift in the cumulative probability plot of sIPSC (G) and mIPSC (H) amplitudes compared to their respective controls (solid lines). K-S test for WT and MCHKO control vs OLZ, p < 0.0001. I, OLZ reduced the amplitude of sIPSC and mIPSC amplitudes in WT and MCHKO MSNs.