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. 2021 Jul 1:12:680494.
doi: 10.3389/fendo.2021.680494. eCollection 2021.

Leptin Receptor Expressing Neurons in the Substantia Nigra Regulate Locomotion, and in The Ventral Tegmental Area Motivation and Feeding

Véronne A J de Vrind  1 , Lisanne J van 't Sant  1 , Annemieke Rozeboom  1 , Mieneke C M Luijendijk-Berg  1 , Azar Omrani  1 , Roger A H Adan  1   2
Affiliations

Leptin Receptor Expressing Neurons in the Substantia Nigra Regulate Locomotion, and in The Ventral Tegmental Area Motivation and Feeding

Véronne A J de Vrind et al. Front Endocrinol (Lausanne). .

Abstract

Leptin is an anorexigenic hormone, important in the regulation of body weight. Leptin plays a role in food reward, feeding, locomotion and anxiety. Leptin receptors (LepR) are expressed in many brain areas, including the midbrain. In most studies that target the midbrain, either all LepR neurons of the midbrain or those of the ventral tegmental area (VTA) were targeted, but the role of substantia nigra (SN) LepR neurons has not been investigated. These studies have reported contradicting results regarding motivational behavior for food reward, feeding and locomotion. Since not all midbrain LepR mediated behaviors can be explained by LepR neurons in the VTA alone, we hypothesized that SN LepR neurons may provide further insight. We first characterized SN LepR and VTA LepR expression, which revealed LepR expression mainly on DA neurons. To further understand the role of midbrain LepR neurons in body weight regulation, we chemogenetically activated VTA LepR or SN LepR neurons in LepR-cre mice and tested for motivational behavior, feeding and locomotion. Activation of VTA LepR neurons in food restricted mice decreased motivation for food reward (p=0.032) and food intake (p=0.020), but not locomotion. In contrast, activation of SN LepR neurons in food restricted mice decreased locomotion (p=0.025), but not motivation for food reward or food intake. Our results provide evidence that VTA LepR and SN LepR neurons serve different functions, i.e. activation of VTA LepR neurons modulated motivation for food reward and feeding, while SN LepR neurons modulated locomotor activity.

Keywords: chemogenetics; dopamine; leptin; midbrain; substantia nigra; ventral tegmental area.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Timeline of experiments.
Figure 2
Figure 2
Immunohistochemistry on LepR-cre x TdTomato mice (n=3). (A) Both SN and VTA expressed LepR neurons (arrows in insets indicate LepR). (B) The majority of LepR neurons co-localized with TH (VTA: 67%, SN: 90%) (C) Of all TH neurons, only 16% in the VTA and 15% in the SN expressed the LepR.
Figure 3
Figure 3
Representative AAV-DIO-ChR2-eYFP tracing of projections from SN LepR neurons in LepR-cre mice (n=4). (A–G) eYFP expression in CPU ++, IPAC ++, STh +, VPM + and VTA +/- to which SN LepR neurons sent detectable projections (++ strong eYFP, + eYFP present, +/- scarce eYFP). In (E–G), eYFP soma and tracks within the SN. B = bregma; CPU, caudate putamen; IPAC, interstitial nucleus of the posterior limb of the anterior commissure; ns, nigrostriatal bundle; SN, substantia nigra; SNc, SN pars compacta; SNr, SN pars reticulate; STh, subthalamic nucleus; VPM, Ventral posteromedial thalamus; VTA, ventral tegmental area.
Figure 4
Figure 4
Expression of AAV-DIO-hM3DGq-mCherry in behavioral animals. Representative example of AAV-DIO-hM3DGq-mCherry expression in (A) VTA LepR-Gq mice (n=6) and (B) SN LepR-Gq mice (n=6). (C, D) Analysis of the location of Gq-mCherry expression showed that the majority of Gq-neurons were found in the targeted region. (E) Analysis of Gq and TH in VTA LepR-Gq mice showed that 57% of VTA Gq neurons co-localized with TH-immunoreactivity. (F) 19% of all VTA TH neurons co-localized with Gq. (G) 96% of SN Gq neurons co-localized with TH-immunoreactivity. (H) Of all SN Th neurons, 18% co-localized with Gq-mCherry.
Figure 5
Figure 5
Active lever presses upon chemogenetic activation of VTA LepR or SN LepR neurons. In ad libitum fed mice, CNO (1.0 and 3.0mg/kg) did not affect active lever presses made in (A) VTA LepR-Gq (n=6) or (B) SN LepR-Gq mice (n=6). (C) Food restricted VTA LepR-Gq mice (n=6) decreased active lever presses after CNO compared to saline injections. (D) CNO did not affect active lever presses in food restricted SN LepR-Gq mice (n=6). Mean ± SEM. *p<0.05, ***p<0.001.
Figure 6
Figure 6
Free 20% sucrose solution consumption upon chemogenetic activation of VTA LepR or SN LepR neurons. CNO (1.0mg/kg) did not affect sucrose solution consumption in ad libitum fed (A) VTA LepR-Gq (n=6) and (B) SN LepR- Gq (n=6) or food restricted (C) VTA LepR-Gq (n=6) and (D) SN LepR-Gq (n=6). Mean ± SEM.
Figure 7
Figure 7
Consumption of chow upon chemogenetic activation of VTA LepR or SN LepR neurons. Chow intake was not affected by CNO (1.0mg/kg) injections in ad libitum fed (A) VTA LepR-Gq (n=6) or (B) SN LepR-Gq mice (n=6). (C) CNO decreased cumulative chow intake in food restricted VTA LepR-Gq mice (n=6). (D) CNO did not affect feeding in food restricted SN LepR-Gq mice (n=6). Mean ± SEM. *p<0.05.
Figure 8
Figure 8
Locomotor activity upon chemogenetic activation of VTA LepR or SN LepR neurons. Locomotion was not affected by CNO (1.0mg/kg) injections in ad libitum fed (A) VTA LepR-Gq (n=6) and (B) SN LepR-Gq (n=6) or (C) food restricted VTA LepR-Gq mice (n=6). (D) CNO decreased locomotion in food restricted SN LepR-Gq mice (n=6). Mean ± SEM. *p<0.05.
Figure 9
Figure 9
Anxiety-like behavior upon chemogenetic activation of VTA LepR or SN LepR neurons. Anxiety-like behavior in the EPM was not affected by CNO (1.0mg/kg) VTA LepR-Gq (n=6) (A) and SN LepR-Gq (n=6) (B) injected mice.
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