And you thought this was a Conspiracy Theory. But, of course it is – except that the conspiracy is on you.
Go On… Get Magnetized as you have nothing to lose but your mind, freedom and liberty – did we mention HUMANITY?
Optogenetic and chemogenetic actuators are critical for deconstructing the neural correlates of behavior. However, these tools have several limitations, including invasive modes of stimulation or slow on/off kinetics. We have overcome these disadvantages by synthesizing a single component, magnetically sensitive actuator, “Magneto,” comprised of the cation channel, TRPV4, fused to the paramagnetic protein, ferritin. We validate non-invasive magnetic control over neuronal activity by demonstrating remote stimulation of cells using in vitro calcium imaging assays, electrophysiological recordings in brain slices, in vivo electrophysiological recordings in the brains of freely moving mice, and behavioral outputs in zebrafish and mice. As proof of concept, we used Magneto to delineate a causal role of striatal dopamine receptor 1 neurons in mediating reward behavior in mice. Together, our results present Magneto as a novel actuator capable of remotely controlling circuits associated with complex animal behaviors.
Opto- and chemogenetic actuators have revealed critical properties of neural networks in normal and pathological states . While both opto- and chemogenetics remotely control neuronal stimulation, optical strategies are limited spatially by poor light penetration into dense tissues and chemogenetic strategies suffer from slow pharmacokinetics that prevent cellular activation on a physiologically relevant timescale. Therefore, there remains a need for next generation actuators that are non-invasive and can respond rapidly and reversibly . Several recent studies have reported transient receptor potential vanilloid 1 (TRPV1) ion channels can be engineered to become sensitive to a combination of radiowaves and magnetothermal heating through coupling to the iron storage protein, ferritin, or to inorganic paramagnetic nanoparticles . While these reagents represent an important advance, they are multicomponent systems (e.g. requiring delivery of nanoparticles and a genetically encoded channel) with possible off-target heating effects. One study has employed non-thermal magnetogenetic control of somatic tissues to regulate blood glucose , but a fully encoded, single component magnetogenetic system has yet to be applied to the nervous system. Here, we have expanded upon these strategies by engineering a magnetogenetic actuator through fusion of the non-selective cation channel, TRPV4 , to the paramagnetic protein, ferritin . We have successfully applied this actuator to the nervous system and validated it using in vitro calcium imaging, brain slice electrophysiology, in vivo electrophysiology, and acute modulation of behavior in freely moving zebrafish and mice.
Design and screen of a novel magnetically sensitive cation channel
To engineer a novel single-component magnetogenetic actuator, we based our design on TRPV4 since it has been reported to respond to pressure . We suspected that, when fused to TRPV4, a paramagnetic protein would enable magnetic torque to tug open the channel to depolarize cells (Supplementary Fig. 1). While we hypothesized that magnetic field dependent activation of TRPV4 would be more facile than using a non-mechanically sensitive ion channel, it may also be formally possible that application of torque to ion channels in general would achieve the same result. Therefore, we developed a small library of 21 proteins consisting of TRPV4 fused to a gene encoding two subunits of the paramagnetic ferritin protein (Supplementary Table 1) . Human embryonic kidney (HEK) 293 cells did not express 18 of the 21 generated chimeric proteins following transient transfection, presumably due to cytotoxicity of the chimeric channels. For the three channels that did express in HEK293 cells, we performed in vitro calcium imaging to determine whether the fusion proteins responded to magnetic
fields. Using the fluorescent calcium-binding dye Fluo-4, we measured calcium transients in response to a ~50 mT magnetic field delivered by an electromagnet (Supplementary Fig. 2). Of the three candidate proteins, we observed detectable calcium transients in response to magnetic stimulation with one fusion protein, consisting of ferritin tethered to a truncated TRPV4 carboxyl-terminus (Δ760–871) (Supplementary Fig. 3). Because the 17±3.5% (mean±SEM) increase in magnetically evoked calcium transients was smaller than expected TRPV4 responses (Supplementary Fig. 3h), we hypothesized that trafficking to the plasma membrane was disrupted , resulting in blunted calcium signaling. We
next optimized the chimeric channel’s subcellular localization by adding a series of subcellular trafficking signals to Magneto akin to the optimization of optogenetic actuators .
Ultimately, we determined that the addition of a plasma membrane trafficking signal enhanced the prototype channel’s membrane expression (Supplementary Fig. 4), and we dubbed this improved channel “Magneto2.0.” We confirmed that HEK293 cells were viable after Magneto2.0 expression (Supplementary Fig. 5) and then measured magnetic field dependent calcium transients produced by Magneto2.0 using the paradigm described in Supplementary Fig. 3. We observed that cells expressing Magneto2.0 (58% transfected cells, n=6 coverslips, n=539 cells) exhibited robust calcium transients approximately 2.5-fold higher than baseline after 50 mT magnetic stimulation with no change in any of the control conditions (Fig. 1a–f). Controls included: (1) cells expressing non-fused TRPV4 and ferritin moieties, (2) unstimulated Magneto2.0 expressing cells, (3) Magneto2.0 expressing cells exposed to the TRP pore blocker ruthenium red (RR), and (4) Magneto2.0 expressing cells in Ca free extracellular media. We observed calcium influx immediately following magnetic stimulation but invariably, maximal calcium fluorescence was observed minutes after magnetic field stimulation of Magneto2.0 expressing cells, which was not observed in any of the above control conditions (Fig. 1g). We found that the delayed calcium response in Magneto2.0+ cells was caused by calcium release from intracellular stores
following magnetically induced depolarization since this secondary response was eliminated following depletion of intracellular calcium stores by thapsigargin, a sarco-endoplasmic reticulum calcium transport ATPase pump inhibitor (Supplementary Fig. 6). However, we sought to determine if the increase in calcium signaling concomitant with magnetic field stimulation was GSK205 sensitive, which would suggest that the signal is TRPV4 dependent . We thus stimulated and quantified the change in calcium fluorescence of mCherry+ Magneto2.0-p2A-mCherry transfected cells during
magnetic field application both in the presence and absence of the specific TRPV4 inhibitor GSK205. We observed a magnetic field dependent calcium increase in the GSK205-untreated Magneto2.0 expressing cells compared to stimulated GSK205-treated cells (two-way ANOVA, p<0.0001) (Fig. 1h). Moreover, 70±5.1% (mean±SEM) of Magneto2.0+ cells responded to magnetic fields (n=3 coverslips,n=58 cells) with an average maximal change in calcium fluorescence of 29±9.8% (mean±SEM) during stimulation compared to only 6.5±0.9% (mean±SEM) for the GSK205-treated population (n=3 coverslips per condition, n=88 GSK205-treated cells, n=57 untreated cells, unpaired two-tailed t-test,t =2.819, p=0.0055). Importantly, all observed changes in calcium fluorescence were noticeably
improved over the poorly trafficked prototype channel (Supplementary Figs. 3, 4a). These data demonstrate that Magneto2.0 is a magnetically sensitive, genetically encoded actuator that can manipulate cellular activity in vitro.
Figure 1 (Remote control of calcium signaling using Magneto2.0)
(a–e) In vitro calcium imaging micrographs of Fluo-4-loaded HEK293 cells before and after 3 pulses of 40–50 mT, 0.1 Hz, 90% duty cycle magnetic stimulation. (f) Quantification of calcium fluorescence fold change in response to the given condition. All experiments treated with magnetic fields except “no magnet” condition. Replicates are shown as individual coverslips equaling n=5 (TRPV4/ferritin), n=3 (No magnet), n=4 (Ruthenium red, (RR)), n=4 (Ca free), n=5 (Magnet) coverslips per condition; total cells analyzed per condition are n=195 (TRPV4/ferritin), n=150 (No magnet), n=148 (RR), n=206 (Ca free), n=396 (Magnet) with n>30 cells analyzed per coverslip. One-way ANOVA, Bonferroni post-test, (F =7.268, p=0.0016). (g) Average temporal kinetics of all cells analzyed within a single coverslip per condition (n=48
(TRPV4/ferritin), n=50 (No magnet), n=45 (RR), n=45 (Ca free), n=102 (Magnet)). Horizontal bar/horseshoe indicates magnetic field application. Two-way ANOVA, Bonferroni post-test, (F =199.1, p<0.0001), *p<0.05 for all time points from 250 s onward compared to “Magnet”. (h) Kinetics of calcium fluorescence fold change within mCherry+ cells in response to magnet in the presence or absence of the TRPV4 inhibitor, GSK205 (10 μM). n=3 coverslips per condition. Data represent all mCherry+ cells analyzed (n=88 GSK205-treated, n=57 untreated). Two-way ANOVA, Bonferroni post-test, (F =23.7, p<0.0001), ***p<0.0001 for all time points from 30 s onward. ***p<0.001, **p<0.01, *p<0.05. Data shown as mean±SEM.
Electrophysiological characterization of Magneto2.0 in the mammalian brain
These preliminary experiments prompted us to precisely determine the temporal kinetics of Magneto2.0 activation since the future utility of Magneto2.0 is contingent on its rapid activation in response to magnetic fields in live tissues. To this end, we generated an adeno-associated virus (AAV) to express Magneto2.0 in mammalian cells under control of the cytomegalovirus (CMV) promoter using the double-floxed inverse open reading frame (DIO) approach (CMV::DIO-Magneto2.0). This strategy enables permanent Cre-dependent expression of a reversed lox site-flanked gene through Cre-lox mediated recombination (Fig. 2a) . We bilaterally co-injected the medial entorhinal cortices (mECs) of WT mice with an AAV1 containing CMV::DIO-Magneto2.0 and an AAV9 carrying Cre recombinase fused to enhanced green fluorescent protein (EGFP) under control of the calcium/calmodulin-dependent protein kinase II alpha (CamKIIα) promoter (CamKIIα::Cre-EGFP), which will express Magneto2.0 in excitatory neurons (Fig. 2b). To test whether Magneto2.0 could elicit action potentials (APs) in neurons from brain slice preparations in response to magnetic fields, we recorded from EGFP+ neurons in the mEC of WT mice doubly transduced with AAVs carrying CMV::DIO-Magneto2.0 and CamKIIα::Cre- EGFP under whole-cell current clamp conditions. Upon application of a ~50 mT static magnetic field delivered by a NdFeB rare earth magnet, neurons in the mEC reliably fired a series of APs akin to spiking behavior evoked by injection of 300 pA of depolarizing current (Fig. 2c, Supplementary Fig. 7a). APs were elicited by both current injection and magnetic fields in 12/12 strongly EGFP+ neurons tested (n=5 mice; n=2 mice excluded due to low EGFP expression). Measurement of time to threshold and time to peak for APs evoked either by current injection or magnetic fields revealed no differences (Supplementary Fig. 7b). Membrane properties, such as resting membrane potential, AP amplitude, upstroke velocity, AP width, and firing threshold were similar between the two stimulation conditions (Supplementary Fig. 7c–g). As controls, we measured that magnetic stimulation initiated APs at a comparable rate to current injection (Supplementary Fig. 8a) and did not cause electrical interference in electrophysiology measurements (Supplementary Fig. 8b). To test if the magnetically evoked firing was due specifically to activation of TRPV4, we bathed brain slices in the selective TRPV4 antagonist GSK205 (n=3 neurons from 3 mice). After a 10-minute incubation with GSK205, magnetic stimulation failed to evoke APs (Fig. 2c, right panel), suggesting that the observed APs were due to Magneto2.0 activation. To determine whether magnetic stimulation affects mEC neurons not expressing Magneto2.0, we magnetically stimulated cells transduced with AAVs delivering CMV::DIOMagneto2.0 and CamKIIα::EGFP, thus preventing Cre-dependent expression of Magneto2.0. We found that stimulation with magnetic fields did not evoke APs in non-Magneto2.0 expressing EGFP+ neurons of the mEC, although these neurons fired spike trains in response to injection with 300 pA of depolarizing current (n=6 neurons from 3 mice) (Fig. 2d, Supplementary Fig. 7h). In sum, we found that only Magneto2.0-expressing neurons of the mEC fired APs in response to magnetic field stimulation, and bath application of GSK205 blocked these responses (Fig. 2e). These data support the notion that activation of Magneto2.0 can rapidly and reversibly depolarize neurons leading to remote control over neural circuit dynamics.
Figure 2 (Electrophysiological characterization of Magneto2.0 in mouse brain slices)
(a) Schematic of viral vector. ITR: inverted terminal repeats; CMV: cytomegalovirus promoter; P: loxP site; 2: lox2272 site. (b) EGFP immunostaining of a WT mouse brain slice showing areas of viral transduction. Hippocampus/entorhinal cortex was doubly transduced with two AAV vectors: AAV1 carrying CMV::DIOMagneto2.0 and AAV9 carrying CaMKIIα::Cre-EGFP. DG: dentate gyrus, sub: subiculum, EC: entorhinal cortex. (c) Magnetically evoked spike train of a current-clamped mEC neuron transduced with CaMKIIα::Cre-EGFP and CMV::DIO-Magneto2.0. Neuron was stimulated with a 50 mT static magnetic field delivered by a permanent magnet. The graded bar represents the magnetic field experienced by neurons during the initiation and cessation of magnetic stimulation as the permanent magnet was brought toward the brain slice using a micromanipulator. Magnetically evoked APs were abolished by bath application of 10 μM GSK205. (d) Sample trace from an EGFP+ current-clamped mEC neuron transduced with CaMKIIα::EGFP and CMV::DIO-Magneto2.0 and thus, not expressing Magneto2.0. No action potentials are elicited in response to magnetic stimulation. (e) Quantification of the number of spikes compared between current injection (n=14 neurons, n=5 mice) and magnetic stimulation (n=12 neurons, n=5 mice) for EGFP+ cells expressing Magneto2.0. No magnetically induced APs are observed during bath application of GSK205 (n=3 neurons, n=3 mice) or when Magneto2.0 is not expressed (300 pA: n=6 neurons, Magnet: n=3 neurons, n=3 mice). All neurons examined are from a total of n=8 mice. Left panel: one-way ANOVA, Bonferroni post-test, (F =4.301, p=0.0243). Right panel: unpaired two-tailed t-test, (t =13.23, p<0.0001). ***p<0.001, *p<0.05, ns: not significant. Data shown as mean±SEM.
Genetically targeted remote magnetic control over zebrafish tactile behaviors
We next began validation of Magneto2.0 function in vivo. We first sought to remotely modulate a simple behavior of the zebrafish, Danio rerio. We transiently expressed Magneto2.0 in Rohon-Beard sensory neurons (5 Magneto2.0+ Rohon-Beard neurons per fish, n=9 fish), using regulatory sequences of the ngn1 promoter . We identified mosaic zebrafish expressing Magneto2.0 in Rohon-Beard neurons by selecting for animals that also expressed a co-injectable fluorescent marker in the heart (Supplementary Fig. 9a). We sought to determine whether magnetic stimulation of zebrafish expressing Magneto2.0 led to an increase in calcium signaling within Rohon-Beard neurons. To this end, we
performed GCaMP imaging of live, 48 hours post-fertilization (hpf) zebrafish larvae expressing Tg(s1020t::Gal4); Tg(UAS::GCaMP3);ngn1::Magneto2.0-p2A-mCherry, which enables detection of activated neurons through the genetically encoded calcium sensor, GCaMP3 , which is expressed in ventral spinal cord neurons . This transgenic combination enables direct visualization of calcium transients in response to magnetic stimulation through dual labeling of GCaMP3+ and mCherry+ Rohon-Beard neurons. We delivered a 50 mT static magnetic field via NdFeB rare earth magnets and
observed an immediate increase in GCaMP3 fluorescence in stimulated Magneto2.0+, mCherry-labeled Rohon-Beard neurons but not in adjacent mCherry- neurons populating the spinal cord (Fig. 3a, Supplementary Fig. 10a). We determined that 17/20 mCherry+ neurons responded above the 6.9±0.15% (mean±SEM) average maximal fluorescence change of control, mCherry- cells (Supplementary Fig. 10a), suggesting that magnetic stimulation in vivo will reliably activate Magneto2.0+ neurons, consistent with both our calcium imaging and slice electrophysiology data. We next tested whether remote activation of Rohon-Beard neurons is sufficient to modulate the behavior of ngn1::Magneto2.0
zebrafish in the presence or absence of magnetic fields. We developed a magnetized behavioral testing arena formed by spacing two NdFeB rare earth magnets 6 mm apart (Supplementary Fig. 10b), which delivered a ten-fold greater magnetic field of ~500 mT to zebrafish larvae than the GCaMP assay. We hypothesized that even if only a few Rohon-Beard neurons were activated by Magneto2.0, the stereotypical escape response would nevertheless induce a coiling behavior, as demonstrated previously . Indeed, in response to a 500 mT magnetic field, groups of 24 to 34 hours post
fertilization (hpf) ngn1::Magneto2.0 expressing zebrafish larvae coiled more frequently compared to those not exposed to a field (Fig. 3b, Supplementary Movies 1–2). In contrast to ngn1::Magneto2.0 fish, which displayed an approximate ten-fold increase in coiling behavior upon magnetic field exposure, there was no observable change in this behavior for either control group—uninjected WT fish or ngn1::TRPV4-p2A-ferritin fish, which bicistronically express independent, unfused TRPV4 and ferritin moieties (Fig. 3c). Consistent with in vitro findings, fish expressing the Magneto prototype
channel under control of the β-actin promoter exhibited a response that was five-fold smaller than that of fish expressing Magneto 2.0 (Supplementary Fig. 9b–d). Finally, we confirmed that Magneto2.0 expression did not disrupt normal peripheral projections of Rohon-Beard neurons by examining red fluorescent protein (RFP) expression in sensory neurons of Tg(isl1::rfp) fish and Tg(isl1::rfp);ngn1::Magneto2.0-IRES-nlsegfp chimeric fish (Supplementary Fig. 10c–f). Together,
these results confirm that Magneto 2.0 is a viable candidate for remotely controlling neuronal activity and animal behavior in vivo.
Figure 3 (Magnetic control over zebrafish tactile behavior in vivo)
(a) Quantification of GCaMP3 fluorescence in mCherry+ Rohon-Beard sensory neurons and mCherryspinal cord neurons in 48 hpf zebrafish larvae expressing ngn1::Magneto2.0-p2A-mCherry. n=20 mCherry+, n=33 mCherry- neurons from 8 stimulation experiments using n=5 zebrafish from 2 independent injection cohorts. Two-way ANOVA, Bonferroni post-test, (F =3.248, p<0.0001). *p<0.05 for all points from 35–55 seconds. (b) Coiling rate of 24–36 hpf ngn1::Magneto2.0 fish. Number of independent experiments for each condition is n=3 (No magnet) and n=6 (Magnet). n=26 (No magnet)
and n=25 (Magnet) fish were used in respective conditions. Unpaired two-tailed t-test, (t =6.152, p=0.0005). (c) Fold change in coiling of fish genotypes aged 24–36 hpf. Number of videos analyzing baseline coiling is n=3 per genotype, number of magnetic stimulation experiments include n=4 (Uninjected), n=4 (ngn1::TRPV4-p2A-ferritin), and n=6 (ngn1::Magneto2.0). Number of fish analyzed shown as (baseline, magnet) for each genotype is: Uninjected: (27, 18), ngn1::TRPV4-p2A-ferritin (17, 21), and ngn1::Magneto2.0 (26, 25). One-way ANOVA, Bonferroni post-test (F =39.01, p<0.0001). Data pooled from 2 independent injection cohorts per genotype. ***p<0.001, *p<0.05. Data shown as
Remote control of mammalian neural activity in freely behaving mice
To determine if Magneto2.0 is capable of controlling mammalian neural activity in vivo, we performed electrophysiology measurements in freely behaving mice transduced with an AAV1 carrying CMV::DIO-Magneto2.0, which expresses Magneto2.0 in a Cre-dependent manner. We aimed to test if Magneto2.0 is capable of rapidly activating a large nucleus deep within the brain, which is more challenging when using optical actuators. To this end, we used mice expressing Cre recombinase under control of the dopamine receptor 1 promoter (Drd1a::Cre), which is expressed in approximately half of
the medium spiny neurons (MSNs) of the striatum . We then transduced striatal neurons of Drd1a::Cre mice with an AAV1 carrying Magneto2.0 and two weeks post-viral injection, we performed extracellular single-unit recordings with tetrode microdrives on Magneto2.0 expressing striatal cells in freely behaving mice and examined the effects of magnetic stimulation on neural firing (Fig. 4a). For this assay, we designed a magnetized chamber (23 cm×4 cm × 18 cm) consisting of NdFeB magnets embedded in the chamber walls (Fig. 4b) and quantified the firing rates of striatal neurons under three conditions: (1) at baseline without magnetic stimulation, (2) during exposure to 50–250 mT magnetic
fields within the chamber, and (3) post-magnetic field exposure. We classified recorded cells into two main groups based on firing rate: slow-spiking (<5 Hz) and fast-spiking (>5 Hz) neurons with mean firing rates of 2.1±0.3 Hz (mean±SEM) and 8.6±0.6 Hz (mean±SEM), previously described as putative MSNs (either D1R+/D2R− or D1R−/D2R+) and GABAergic interneurons (D1R−), respectively. Exposure of these mice to magnetic fields produced a 43.8±20.3% increase in the overall firing rate of slow-spiking putative MSNs (Fig. 4c–e). Importantly, the firing rate of putative GABAergic interneurons remained constant (Fig. 4c–d). Subsequent to magnetic stimulation, 66.7% of putative
MSNs returned to baseline firing rates, while the putative interneuron firing rate again remained at baseline (Fig. 4f). Finally, we observed an increase in the firing rate of slow-spiking, but not fastspiking, neurons of the striatum following systemic administration of the D1R agonist, SKF81297 (Supplementary Fig. 11a), suggesting that the D1R+ population responsive to magnetic fields are indeed slow-spiking neurons. Together, these data demonstrate that Magneto2.0 is capable of controlling neural firing in deep brain regions in response to magnetic fields.
Figure 4 (Magnetogenetic control of the mammalian nervous system in vivo)
(a) Representation of magnetic stimulation and recording of D1R-expressing cells in the striatum of Drd1a::Cre mice. Solid lines indicate electrode placement from 5 mice; dashed circle indicates approximate injection area. (b) Cartoon of magnetized testing chamber, rare earth magnets (gray bars) are embedded in the walls, “B” represents magnetic field, magnetic field strength shown as gradient. (c) Quantification of single unit average firing rate during magnetic field exposure in freely behaving mice. n=51 <5 Hz neurons, n=81 >5 Hz neurons from 5 mice (n=66, n=30, n=25, n=7, n=4 cells from each mouse). Unpaired two-tailed t-test, (t =3.210, p=0.0017). (d) Proportion of cells firing >5% over baseline
during magnet exposure. (e) Standard score (z-score) over time for <5 Hz MSNs in d that fired >5% (red,
n=23) vs. <5% (black, n=28). Two-way ANOVA (F =210.9, p<0.0001). Gray box represents stimulation in magnetized chamber. Dashed line shows baseline of no change. (f) Proportion of cells firing >5% over baseline post-magnet exposure. Data are shown as mean±SEM. ***p<0.001, **p<0.01.
Remote magnetogenetic control of D1R-mediated striatal reward valence
Ultimately, we sought to determine whether Magneto2.0 dependent control of neural activity in vivo could translate to control over complex mammalian reward behaviors regulated by dopamine signaling . While optogenetic studies have implicated the dopaminergic signaling axis in causally mediating reward behavior , it is unclear whether activation of postsynaptic D1R+ neurons is sufficient for controlling this effect. For instance, optogenetic stimulation of one subset of striatal D1R+ neurons is not causally responsible for induction of conditioned place preference (CPP) . Conversely,
studies using systemic pharmacological manipulations with D1R agonists confirm that activation of D1R+ neurons is sufficient to evoke CPP , suggesting that broadly activating D1R+ neurons may cause reinforcing behaviors. Optogenetic techniques are intrinsically limited in the number of neurons that can be activated simultaneously via fiberoptic implants and pharmacological approaches lack genetic specificity. However, a magnetogenetic paradigm circumvents both obstacles simultaneously allowing resolution of this discrepancy with cell-type specificity and a real time behavioral output. We tested the sufficiency of D1R+ neurons in eliciting reward conditioning by unilaterally injecting the striata of WT and Drd1a::Cre mice with an AAV1 carrying CMV::DIO-Magneto2.0 and subjecting the mice to a real time place preference (RTPP) assay where they could choose between a magnetized arm, lined with eight permanent NdFeB magnets delivering a magnetic field gradient of 250–50 mT, and a non-magnetized arm (Fig. 5a). We observed that Magneto2.0 expressing Drd1a::Cre mice showed a significant preference for the magnetized arm of the RTPP chamber in contrast to WT mice (one-way ANOVA, p=0.0152), which exhibited no preference (Fig. 5b–e). Moreover, removal of the magnets from the chamber eliminated the preference of Magneto2.0 expressing Drd1a::Cre mice for either arm, a response identical to WT mice (Fig. 5c), demonstrating that RTPP is dependent on D1R stimulation. As a control, we measured no differences in overall locomotion between unilaterally injected WT and Drd1a::Cre mice using a modified open field assay (Supplementary Fig. 11b–c). These data show: (1) that broad activation of D1R+ neurons of the striatum is sufficient to control reward salience and (2) that Magneto2.0 can be used for remote control of complex mammalian behaviors mediated by deep brain nuclei in freely moving mice.