Research Articles: Systems/Circuits Dynamic impairment of olfactory behavior and signaling mediated by an olfactory corticofugal system https://doi.org/10.1523/JNEUROSCI.2667-19.2020 Cite as: J. Neurosci 2020; 10.1523/JNEUROSCI.2667-19.2020 Received: 10 November 2019 Revised: 30 June 2020 Accepted: 5 July 2020 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 © 2020 the authors 1 Journal Section 2 Systems/Circuits 3 4 Title 5 Dynamic impairment of olfactory behavior and signaling mediated by an olfactory 6 corticofugal system 7 8 Abbreviated title 9 Dynamic inhibition of olfactory sensory responses 10 11 Authors 12 Renata Medinaceli Quintela1, Jennifer Bauer1, Lutz Wallhorn1, Kim Le1, Daniela Brunert1, 13 Markus Rothermel1 14 15 1. Department of Chemosensation, AG Neuromodulation, Institute for Biology II, RWTH 16 Aachen University, Aachen 52074, Germany 17 18 19 Corresponding author 20 Markus Rothermel 21 RWTH Aachen University 22 D-52074 Aachen 23 [email protected] 24 25 Number of figures: 8 26 Number of tables: 1 27 Number of pages: 47 28 Abstract: 205 words 29 Introduction: 662 words 30 Discussion: 1949 words 31 32 Conflicts of interest 33 None 34 35 36 37 Acknowledgments 38 We thank Scott W. Rogers and Petr Tvrdik for kindly providing the Chrna7-Cre mice. 39 Chrna7-Cre mice were generated with funding from NIH AG017517 to S. Rogers. We thank 40 the technical workshop at the institute for biology II, RWTH Aachen for excellent technical 41 support. We thank Matt Wachowiak, Jeremy C. McIntyre, and all members of the M.R. lab 42 for helpful discussion and comments on the manuscript. Initial data were acquired in Utah. 43 We thank Drs. Looger, Akerboom, and Kim and the Genetically Encoded Calcium Indicator 44 (GECI) Project at Janelia Farm Research Campus in collaboration with Penn Vector Core for 45 providing with GCaMP-expressing viruses. This work was supported by funding from DFG 46 (RO4046/2-1 and /2-2, Emmy Noether Program [to MR] and the Research Training Group 47 2416 “MultiSenses – MultiScales: Novel approaches to decipher neural processing in 48 multisensory integration” 368482240/GRK2416) and by a grant from the Interdisciplinary 49 Centre for Clinical Research within the faculty of Medicine at the RWTH Aachen University 50 (IZKF TN1-7 532007). 51 1 52 Abstract 53 Processing of olfactory information is modulated by centrifugal projections from 54 cortical areas, yet their behavioral relevance and underlying neural mechanisms remain 55 unclear in most cases. The anterior olfactory nucleus (AON) is part of the olfactory cortex and 56 its extensive connections to multiple upstream and downstream brain centers place it in a 57 prime position to modulate early sensory information in the olfactory system. Here, we show 58 that optogenetic activation of AON neurons in awake male and female mice was not 59 perceived as an odorant equivalent cue. However, AON activation during odorant 60 presentation reliably suppressed behavioral odor responses. This AON mediated effect was 61 fast and constant across odors and concentrations. Likewise, activation of glutamatergic AON 62 projections to the olfactory bulb (OB) transiently inhibited the excitability of mitral/tufted 63 cells (MTCs) that relay olfactory input to the cortex. Single-unit MTC recordings revealed 64 that optogenetic activation of glutamatergic AON terminals in the OB transiently decreased 65 sensory-evoked MTC spiking, regardless of the strength or polarity of the sensory response. 66 The reduction in MTC firing during optogenetic stimulation was confirmed in recordings in 67 awake mice. These findings suggest that glutamatergic AON projections to the OB impede 68 early olfactory signaling by inhibiting OB output neurons thereby dynamically gating sensory 69 throughput to the cortex. 70 71 Significance Statement 72 The anterior olfactory nucleus (AON) as an olfactory information processing area 73 sends extensive projections to multiple brain centers but the behavioral consequences of its 74 activation have been scarcely investigated. Using behavioral tests in combination with 75 optogenetic manipulation we show that in contrast to what has been suggested previously, the 76 AON does not seem to form odor percepts but instead suppresses behavioral odor responses 2 77 across odorants and concentrations. Furthermore, this study shows that AON activation 78 inhibits olfactory bulb output neurons in both anesthetized as well as awake mice, pointing to 79 a potential mechanism by which the olfactory cortex can actively and dynamically gate 80 sensory throughput to higher brain centers. 3 81 Introduction 82 The ability to perceive external information via sensory systems is crucial for an 83 animal to navigate and survive in a complex environment. In a classical view, the brain 84 processes sensory information solely based on a hierarchical organization where sensory 85 information is shaped and refined by subsequent processing steps. However, to guarantee 86 appropriate, flexible, and fast reactions in a rapidly changing environment, it is beneficial to 87 implement additional mechanisms that modulate information in a situation-dependent fashion. 88 One way to do so are cortical top-down projections, where sensory information is received 89 from sensory cortices, and processed information is then transmitted to downstream centers to 90 modulate incoming sensory signals. Understanding the neural mechanisms underlying sensory 91 perception thus requires information on the neural circuits involved in both bottom-up and 92 top-down mechanisms. 93 One prominent center of cortical top-down projections in olfaction is the anterior 94 olfactory nucleus (AON), an olfactory cortical area located in the forebrain just caudally of 95 the olfactory bulb (OB), the first relay station of olfactory signals within the brain. The AON 96 can be divided into two distinct zones, pars externa, a thin ring of cells in the rostral part of 97 the AON, and pars principalis containing the majority of AON cells (Valverde et al., 1989; 98 Brunjes et al., 2005). Its extensive connectivity with primary and secondary processing 99 centers (see (Brunjes et al., 2005)) and its position as both a “bottom-up” relay of ascending 100 sensory input from the OB and a source of “top-down” input to the OB render the AON an 101 interesting model system for investigating higher-order olfactory processing and the interplay 102 of ascending and descending information. 103 The AON is the largest source of cortical projections to the OB (Carson, 1984; Shipley 104 and Adamek, 1984). AON-derived axons have been shown to project to multiple layers of the 105 OB (Reyher et al., 1988; Padmanabhan et al., 2016; Wen et al., 2019). This includes the 106 granule cell layer which contains the majority of inhibitory interneurons of the OB, as well as 4 107 the layers containing the output neurons of the OB, the external plexiform, and the mitral cells 108 layer. Furthermore, AON projections are bilateral, i.e. the AON does not only send axons to 109 the ipsilateral but also, via the anterior commissure, to the contralateral OB (Brunjes et al., 110 2005; Illig and Eudy, 2009; Wen et al., 2019). Similar to cortical back projections from 111 piriform cortex (Boyd et al., 2015; Otazu et al., 2015), the AON was shown to send sensory- 112 evoked feedback to the OB (Rothermel and Wachowiak, 2014). 113 The AON has been implicated in a range of different functions, including serving as 114 the first site of integrated odor percept formation (Haberly, 2001; Wilson and Sullivan, 2011), 115 olfactory memory (Haberly, 2001; Aqrabawi and Kim, 2018b, 2020; Levinson et al., 2020), 116 social interaction (Wacker et al., 2011; Oettl et al., 2016; Wang et al., 2020), controlling food 117 intake (Soria-Gomez et al., 2014), and integrating activity within and between the two OBs 118 (Schoenfeld and Macrides, 1984; Lei et al., 2006; Yan et al., 2008; Kikuta et al., 2010; 119 Esquivelzeta Rabell et al., 2017; Grobman et al., 2018). Despite this wide variety of proposed 120 functions the exact role of centrifugal AON projections in modulating ongoing OB activity 121 remains poorly characterized. Only a few studies have investigated the influence of 122 centrifugal AON projections on OB circuit function (Markopoulos et al., 2012; Oettl et al., 123 2016; Grobman et al., 2018); demonstrating that AON inputs can depolarize as well as inhibit 124 mitral/tufted cells (MTC). 125 In the present study, we used optogenetic AON stimulation to decipher AON effects 126 on odor related behavior. Whereas AON stimulation was not perceived as an odor equivalent 127 cue, AON activation during odorant presentation reliably suppressed behavioral odor 128 responses. This effect was constant across odors and concentrations. Optical AON stimulation 129 in anesthetized as well as in awake mice resulted in a substantial decrease in MTC spiking 130 during sensory stimulus presentation matching the behavioral results. These findings support 131 the hypothesis that the AON acts as a strong regulator of olfactory information transmitted to 132 higher brain areas. 5 133 134 Materials and Methods 135 Animals strain and care 136 We used a mouse line (Chrna7-Cre, kindly provided by S. Rogers and P. Tvrdik, 137 University of Utah) in which an IRES-Cre bi-cistronic cassette was introduced into the 138 3’noncoding region of the cholinergic nicotinic receptor alpha7 (Chrna7) (Rogers and 139 Gahring, 2012; Rogers et al., 2012a; Rogers et al., 2012b; Gahring et al., 2013). Animals of 140 either sex were used. Animals were housed under standard conditions in ventilated racks.