Adolescent neurostimulation of dopamine circuit reverses genetic deficits in frontal cortex function

Reviewer #1 (Public Review):

The current study was designed to test the hypothesis that neural circuit plasticity during adolescence can be targeted to restore cortical function under conditions of developmental disruptions that are relevant to psychiatric disorders. Specifically, the authors targeted the mesofrontal cortical dopamine system in 2 genetic mouse models of schizophrenia and performed optical recordings in combination with behavior and chemogenetic manipulations. Major findings and strengths include that stimulation of frontal dopaminergic projections in a limited adolescent time window can stably reverse defects in cortical neuronal activity and cognitive control in adulthood in 2 genetic mouse models of psychiatric disorders. While the precise postsynaptic mechanisms underlying the positive impact of adolescent mesofrontal dopamine stimulation were not address, another strength of this study is that the authors performed key manipulations using age and dose/intensity as dependent variables to show that the level of neural circuit activation during adolescence follows an inverted U-shape pattern. Collectively, this is a well-design study with many strengths and novel findings that are likely to positively impact a widespread of disciplines within the biological psychiatry and neuroscience field.

Reviewer #2 (Public Review):

The manuscript by Mastwal and colleagues explores how transient adolescent stimulation of ventral midbrain neurons that project to the frontal cortex may help to improve performance on certain memory tasks. The manuscript provides an interesting set of observations that DREADD-based activation over only 3 days during adolescence provides a fast-acting and long-lasting improvement in performance on Y-maze spontaneous alternation as well as aspects of neuronal function as assessed using in vivo imaging methods. While interesting, there are several weaknesses. First and foremost, it is not clear that the effects the authors are observing are mediated by dopamine. It has been clearly documented that the DAT-Cre line provides a better representation of midbrain dopamine cells in the mouse, particularly near the midline of the ventral midbrain (Lammel et al., Neuron 2015). This is precisely where the cells that project to the frontal cortex are located. Therefore, the selection of TH-Cre is problematic. It is very likely that the authors are labeling a substantial number of non-dopaminergic cells.

Reviewer #3 (Public Review):

In this manuscript, the authors use behavior, calcium imaging, and circuit modulation (DREADDs etc) to assess dopamine regulation of prefrontal cortical circuits in the mouse. The authors have previously established that activation of dopamine inputs to prefrontal cortex during adolescence can drive increases in mPFC DA bouton number and enhanced mPFC activity in WT mice. Here the authors use two mouse models - one with a reporter replacing the Arc gene, and another with knockout of the schizophrenia-associated gene Disc1, both of which are thought to have reduced prefrontal cortical activity. First they trained mice on a Y-maze and showed impaired performance in the Arc knockout. Then they demonstrated selective disruption of neuronal firing with calcium imaging at the time of the decision in the task. The Arc mice were found to have reduced dopamine bouton density, and adolescent activation of the DA neurons corrected this as well as the PFC firing and the behavior. Similar data were shown in the Disc1 KO. The data are well controlled and the authors use a number of leading edge methods.

Author Response:

First and foremost, we would like to thank all the editors and reviewers for their thoughtful and thorough evaluations of our manuscript. We greatly appreciate their assessment about the novelty and strength in this study and will revise the manuscript according to their recommendations. Here we offer a provisional response to Reviewer 2 to clarify our rationale for using TH-Cre rather than DAT-Cre mice in our study of frontal cortical dopaminergic projections.

We agree with Review 2 that the DAT-Cre line can provide specific labeling of midbrain dopamine neurons projecting to the striatum, as discussed in the cited study (Lammel et al., 2015). But unfortunately, mesocortical dopamine neurons in the VTA are known to express very little DAT (Lammel et al., 2008; Li, Qi, Yamaguchi, Wang, & Morales, 2013; Sesack, Hawrylak, Matus, Guido, & Levey, 1998). This limitation in the use of the DAT-Cre line to target mesocortical dopamine neurons has been acknowledged in the cited publication (Lammel et al., 2015). It is an issue we have also observed when testing the DAT-Cre line in our lab. Additionally, and interestingly, recent extensive evaluation of the DAT-Cre line reported ectopic labeling of multiple non-dopaminergic neuronal populations (Papathanou, Dumas, Pettersson, Olson, & Wallen-Mackenzie, 2019; Soden et al., 2016; Stagkourakis et al., 2018). Our own evaluation of the DAT-Cre line’s utility for cortical imaging also captured sporadic ectopic labeling of cortical cell somas.

Because mesocortical dopamine neurons have stronger TH expression than DAT (Lammel et al., 2008; Lammel et al., 2015; Li et al., 2013; Sesack et al., 1998), TH-Cre lines have been frequently used to study the mesocortical pathway (Ellwood et al., 2017; Gunaydin et al., 2014; Lammel et al., 2012; Lohani, Martig, Deisseroth, Witten, & Moghaddam, 2019; Vander Weele et al., 2018). While TH-Cre expression itself is not restricted to dopaminergic neurons, we targeted our viral injections to the VTA and optogenetic stimulation to the cortical dopaminergic projection target area (Patriarchi et al., 2018) to specifically modulate mesocortical dopaminergic axons. In addition, we tested D1 antagonist’s effects in our manipulations. Although we targeted dopamine neurons in our adolescent stimulation, the final behavioral outcome likely includes contributions from co-released neurotransmitters and non-dopaminergic neurons via network effects. We will revise our discussion and methods sections to clarify these points of interest. Additionally, we will provide DAT-Cre images in the revised supplementary materials to further explain our choice of the TH-Cre line rather than the DAT-Cre line for our study.

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