Research Articles: Systems/Circuits Superior colliculus controls the activity of the rostromedial tegmental nuclei in an asymmetrical manner https://doi.org/10.1523/JNEUROSCI.1556-20.2021 Cite as: J. Neurosci 2021; 10.1523/JNEUROSCI.1556-20.2021 Received: 25 August 2020 Revised: 12 January 2021 Accepted: 20 February 2021 This Early Release article has been peer-reviewed and accepted, but has not been through the composition and copyediting processes. The final version may differ slightly in style or formatting and will contain links to any extended data. Alerts: Sign up at www.jneurosci.org/alerts to receive customized email alerts when the fully formatted version of this article is published. Copyright © 2021 Pradel et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license, which permits unrestricted use, distribution and reproduction in any medium provided that the original work is properly attributed. 1 Title 2 Superior colliculus controls the activity of the rostromedial tegmental nuclei in an asymmetrical 3 manner 4 Abbreviated title 5 SC controls the RMTg in an asymmetrical manner 6 Authors names and affiliations 7 Kamil Pradel, MSc1, Gniewosz Drwięga, MSc1, Tomasz Błasiak, PhD1 8 1 Department of Neurophysiology and Chronobiology, Institute of Zoology and Biomedical Research, 9 Jagiellonian University, 9 Gronostajowa street, 30-387 Krakow, Poland 10 Corresponding authors’ email addresses: Kamil Pradel: [email protected], Tomasz Błasiak: 11 [email protected] 12 Number of pages: 29 13 Number of: 14 x figures: 9 15 x tables: 0 16 x multimedia: 0 17 x 3D models: 0 18 x extended data figures: 0 19 x extended data tables: 0 20 Number of words for: 21 x abstract: 239 22 x introduction: 623 23 x discussion: 1499 24 Conflict of interest statement 25 The authors declare no competing financial interests. 26 Acknowledgments 27 This work has been supported by the Polish National Science Center grant UMO- 28 2017/27/N/NZ4/00785, awarded to Kamil Pradel, and the statutory funds of the Institute of Zoology 29 and Biomedical Research, Jagiellonian University. The open-access publication of this article was 30 funded by the Priority Research Area BioS under the program “Excellence Initiative – Research 31 University” at the Jagiellonian University in Krakow. 32 Abstract 33 Dopaminergic (DA) neurons of the midbrain are involved in controlling animals’ orienting and 34 approach toward relevant external stimuli. The firing of DA neurons is regulated by many brain 35 structures; however, the sensory input is provided predominantly by the ipsilateral superior colliculus 36 (SC). It is suggested that SC also innervates the contralateral rostromedial tegmental nucleus 37 (RMTg)—the main inhibitory input to DA neurons. Therefore, this study aimed to describe the 38 physiology and anatomy of the SC-RMTg pathway. To investigate the anatomical connections within 39 the circuit of interest, anterograde, retrograde, and transsynaptic tract-tracing studies were performed 40 on male Sprague-Dawley rats. We have observed that RMTg is monosynaptically innervated 41 predominantly by the lateral parts of the intermediate layer of the contralateral SC. To study the 42 physiology of this neuronal pathway, we conducted in vivo electrophysiological experiments combined 43 with optogenetics; the activity of RMTg neurons was recorded using silicon probes while either 44 contralateral or ipsilateral SC was optogenetically stimulated. Obtained results revealed that activation 45 of the contralateral SC excites majority of RMTg neurons, while stimulation of the ipsilateral SC 46 evokes similar proportions of excitatory or inhibitory responses. Consequently, single-unit recordings 47 showed that activation of RMTg neurons innervated by the contralateral SC, or stimulation of 48 contralateral SC-originating axon terminals within the RMTg, inhibits midbrain DA neurons. Together, 49 the anatomy and physiology of the discovered brain circuit suggest its involvement in the animals’ 50 orienting and motivation-driven locomotion based on the direction of external sensory stimuli. 51 Significance statement 52 Dopaminergic neurons are the target of predominantly ipsilateral, excitatory innervation originating 53 from the superior colliculus (SC). However, we demonstrate in our study that SC inhibits dopaminergic 54 neurons’ activity on the contralateral side of the brain via the rostromedial tegmental nucleus. In this 55 way, sensory information received by the animal from one hemifield could induce opposite effects on 56 both sides of the dopaminergic system. It was shown that the side to which an animal directs its 57 behavior is a manifestation of asymmetry in dopamine release between left and right striatum. Animals 58 tend to move oppositely to the hemisphere with higher striatal dopamine concentration. This explains 59 how the above-described circuit might guide animals’ behavior according to the direction of incoming 60 sensory stimuli. 61 2 62 Introduction 63 The role of dopaminergic neurons located in the midbrain (ventral tegmental area, or VTA, and 64 substantia nigra pars compacta, or SNc) in control of motor functions, learning, and reward-related 65 behaviors is well established (Mogenson et al., 1980; Wise, 2004; Baik, 2013). They drive the 66 organism to choose proper motor actions in given circumstances in order to maximize survival. Such 67 action-gating property of dopaminergic neurons requires them to integrate wide range of information, 68 of both external and internal origin. Such a variety of information is provided by the prolific 69 innervation from different brain areas (Watabe-Uchida et al., 2012). Sensorial information is most 70 probably delivered by the superior colliculus (SC), which is a brain region processing sensory 71 information from the contralatearal side of the body (May, 2006; May et al., 2009; Redgrave et al., 72 2010). Notably, the function of intermediate and deep layers of the SC in motor guidance, driving 73 object-oriented eye (saccades), head, body, and limb movements is well documented (Gandhi & 74 Katnani, 2011). In contrast, superficial layers of the SC are mostly visual, involved in rapid detection of 75 the onset of sensory stimuli and further relaying this information to the deeper SC layers. 76 It has been shown that SC innervates midbrain dopaminergic neurons (Comoli et al., 2003; Coizet et 77 al., 2007; May et al., 2009; Yetnikoff et al., 2015), and latency of SC’s reaction to visual stimuli is 78 always shorter than that of midbrain dopaminergic neurons (Redgrave & Gurney, 2006). Phasic activity 79 of dopaminergic neurons evoked by the visual stimuli is diminished by the lesion of the ipsilateral SC 80 (Comoli et al., 2003). Accordingly, pharmacological disinhibition of the SC increased visually evoked 81 (Comoli et al., 2003; Bertram et al., 2014) and spontaneous (Coizet et al., 2003) activity of midbrain 82 dopaminergic neurons, as well as dopamine release in the striatum (Dommett et al., 2005). Moreover, 83 when the SC of monkeys with lesioned primary visual was pharmacologically blocked, it was only then 84 that the monkeys lost the ability to perform anticipatory behaviors based on the reward-associated cues; 85 furthermore, dopaminergic neurons stopped responding to these cues (Takakuwa et al., 2017). 86 Additionally, it was demonstrated recently that the SC to VTA pathway is important for orienting 87 behavior during interaction with a conspecific, as well as mediating visually evoked innate defensive 88 responses (Prévost-Solié et al., 2019; Zhou et al., 2019). 89 Studies mentioned above focus on the innervation of dopaminergic neurons descending from the 90 ipsilateral SC. Nevertheless, anatomical studies strongly suggest that SC also sends projections to the 91 contralaterally located rostromedial tegmental nucleus (RMTg), which constitutes the main inhibitory 92 input to the midbrain dopaminergic neurons (Jhou et al., 2009a, 2009b; Kaufling et al., 2009; Bourdy 93 and Barrot, 2012; Yetnikoff et al., 2015). Given that SC innervates dopaminergic neurons located 94 ipsilaterally and RMTg neurons located contralaterally, it can be hypothesized that the SC controls the 95 activity of dopaminergic neurons located in both hemispheres in an opposite manner (Fig. 1). 3 96 Notably, such brain wiring might have behavioral consequences, as the side toward which the animal 97 directs its behavior is the manifestation of the difference in the activity of the left and right 98 dopaminergic system. Body movements and behaviors are performed toward the hemisphere with 99 lower dopamine release and away from the hemisphere with higher dopamine release (Arbuthnott & 100 Crow, 1971; Iwamoto et al., 1976; Joyce et al., 1981; Glick et al., 1988; Bourdy et al., 2014; 101 Molochnikov & Cohen, 2014). 102 Therefore, SC, via contralateral RMTg, might contribute to the lateralization of an animal’s movement, 103 based on the direction of incoming sensory stimuli. In this anatomical and electrophysiological study, 104 we aimed to describe the innervation descending from the SC to the RMTg and to determine how this 105 neuronal pathway influences the dopaminergic system. To date, this brain circuit has not been 106 investigated. 107 Materials and methods 108 1) Subjects 109 Male Sprague Dawley rats (280–350 g) were acquired from the Institute of Zoology and Biomedical 110 Research, Jagiellonian University (Krakow, Poland) breeding facility. Animals