Young-Cho Kim

Background Dopamine is a powerful neuromodulator of diverse brain functions, including movement, motivation, reward, and cognition. D1-type dopamine receptors (D1DRs) are the most prevalently expressed dopamine receptors in the brain. Neurons expressing D1DRs are heterogeneous and involve several subpopulations. Studying these neurons has been limited by current animal models, especially when considering their integration with conditional or intersectional genetic tools. New method To address this limitation, we developed a novel Drd1-P2A-Flpo (Drd1-Flpo) mouse line in which the Flpo gene was knocked in immediately after the Drd1 gene using CRISPR-Cas9. We validated the Drd1-Flpo line by confirming Flp expression and functionality specific to D1DR+ neurons. Comparison with existing methods: The Drd1-Flpo line is useful resource for studying subpopulation of D1DR+ neurons with intersectional genetic tools. Conclusions We demonstrated brain-wide GFP expression driven by Drd1-Flpo, suggesting that this mouse line may be useful for comprehensive anatomical and functional studies in many brain regions. The Drd1-Flpo model will advance the study of dopaminergic signaling by providing a new tool for investigating the diverse roles of D1DR+ neurons and their subpopulations in brain disease. Significance Statement The roles of dopamine in the brain are mediated by dopamine receptors. D1-type dopamine receptors (D1DRs) and D1DR-expressing (D1DR+) neurons play important roles in various brain functions. We generated a Drd1-Flpo mouse line that expresses Flp recombinase in D1DR+ neurons. This novel Drd1-Flpo mouse facilitates investigation of specific roles of D1DR+ neurons in various brain areas including the striatum, frontal cortex, and cerebellum, and it provides an alternative to existing Drd1-Cre mice. In addition, the Drd1-Flpo mouse line provides a tool for intersectional genetic studies, when used with existing transgenic Cre lines. The Drd1-Flpo mouse line can help unravel the specific contributions of D1DR+ neuron subpopulations to brain function and dysfunction.

Behavioral flexibility requires the prefrontal cortex and striatum, but it is unclear if these structures play similar or distinct roles in adapting to novel circumstances. Here, we investigate neuronal ensembles in the medial frontal cortex (MFC) and the dorsomedial striatum (DMS) during one form of behavioral flexibility: learning a new temporal interval. We studied corticostriatal neuronal activity as rodents trained to respond after a 12-s fixed interval (FI12) learned to respond at a shorter 3-s fixed interval (FI3). On FI12 trials, we found that a key form of temporal processing—time-related ramping activity—decreased in the MFC but did not change in the DMS as animals learned to respond at a shorter interval. However, while MFC and DMS ramping was stable with successive days of two-interval performance, temporal decoding by DMS ensembles improved on FI3 trials. Finally, when comparing FI12 versus FI3 trials, we found that more DMS neurons than MFC neurons exhibited differential interval-related activity early in two-interval performance. These data suggest that the MFC and DMS play distinct roles during temporal learning and provide insight into corticostriatal circuits.

Arousal from sleep in response to CO₂is a critical protective phenomenon. Dysregulation of CO₂-induced arousal contributes to morbidity and mortality from prevalent diseases, such as obstructive sleep apnea and sudden infant death syndrome. Despite the critical nature of this protective reflex, the precise mechanism for CO₂-induced arousal is unknown. Because CO₂is a major regulator of breathing, prevailing theories suggest that activation of respiratory chemo- and mechano-sensors is required for CO₂-induced arousal. However, populations of neurons that are not involved in the regulation of breathing are also chemosensitive. Among these are serotonin (5-HT) neurons in the dorsal raphe nucleus (DRN) that comprise a component of the ascending arousal system. We hypothesized that direct stimulation of these neurons with CO₂could cause arousal from sleep independently of enhancing breathing. Dialysis of CO₂-rich acidified solution into DRN, but not medullary raphe responsible for modulating breathing, caused arousal from sleep. Arousal was lost in mice with a genetic absence of 5-HT neurons, and with acute pharmacological or optogenetic inactivation of DRN 5-HT neurons. Here we demonstrate that CO₂can cause arousal from sleep directly, without requiring enhancement of breathing, and that chemosensitive 5-HT neurons in the DRN critically mediate this arousal. Better understanding mechanisms underlying this protective reflex may lead to interventions to reduce disease-associated morbidity and mortality.SIGNIFICANCE STATEMENTAlthough CO₂-induced arousal is critical to a number of diseases, the specific mechanism is not well understood. We previously demonstrated that serotonin (5-HT) neurons are important for CO₂-induced arousal, as mice without 5-HT neurons do not arouse to CO₂. Many have interpreted this to mean that medullary 5-HT neurons that regulate breathing are important in this arousal mechanism. Here we found that direct application of CO₂-rich aCSF to the dorsal raphe nucleus, but not the medullary raphe, causes arousal from sleep, and that this arousal was lost with genetic ablation or acute inhibition of 5-HT neurons. We propose that 5-HT neurons in the dorsal raphe nucleus can be activated directly by CO₂to cause arousal independently of respiratory activation.

The subthalamic nucleus is a key site controlling motor function in humans. Deep brain stimulation of the subthalamic nucleus can improve movements in patients with Parkinson’s disease; however, for unclear reasons, it can also have cognitive effects. Here, we show that the human subthalamic nucleus is monosynaptically connected with cognitive brain areas such as the prefrontal cortex. Single neurons and field potentials in the subthalamic nucleus are modulated during cognitive processing and are coherent with 4-Hz oscillations in medial prefrontal cortex. These data predict that low-frequency deep brain stimulation may alleviate cognitive deficits in Parkinson’s disease patients. In line with this idea, we found that novel 4-Hz deep brain stimulation of the subthalamic nucleus improved cognitive performance. These data support a role for the human hyperdirect pathway in cognitive control, which could have relevance for brain-stimulation therapies aimed at cognitive symptoms of human brain disease.

Considerable evidence has shown that prefrontal neurons expressing D1-type dopamine receptors (D1DRs) are critical for working memory, flexibility, and timing. This line of work predicts that frontal neurons expressing D1DRs mediate cognitive processing. During timing tasks, one form this cognitive processing might take is time-dependent ramping activity — monotonic changes in firing rate over time. Thus, we hypothesized the prefrontal D1DR+ neurons would strongly exhibited time-dependent ramping during interval timing. We tested this idea using an interval-timing task in which we used optogenetics to tag D1DR+ neurons in the mouse medial frontal cortex (MFC). While 23% of MFC D1DR+ neurons exhibited ramping, this was significantly less than untagged MFC D1DR+ neurons. By contrast, MFC D1DR+ neurons had strong delta-frequency (1-4 Hz) coherence with other MFC ramping neurons. This coherence was phase-locked to cue onset and was strongest early in the interval. To test the significance of these interactions, we optogenetically stimulated MFC D1DR+ neurons early vs. late in the interval. We found that 2-Hz stimulation early in the interval was particularly effective in rescuing timing-related behavioral performance deficits in dopamine-depleted animals. These findings provide insight into MFC networks and have relevance for disorders such as Parkinson’s disease and schizophrenia. Significance Statement: Prefrontal D1DRs are involved in cognitive processing and cognitive dysfunction in human diseases such as Parkinson’s disease and schizophrenia. We use optogenetics to identify these neurons, as well as neurons that are putatively connected to MFC D1DR+ neurons. We study these neurons in detail during an elementary cognitive task. These data could have relevance for cognitive deficits for Parkinson’s disease, schizophrenia, and other diseases involving frontal dopamine.

Although frontostriatal circuits are critical for the temporal control of action, how time is encoded in frontostriatal circuits is unknown. We recorded from frontal and striatal neurons while rats engaged in interval timing, an elementary cognitive function that engages both areas. We report four main results. First, “ramping” activity, a monotonic change in neuronal firing rate across time, is observed throughout frontostriatal ensembles. Second, frontostriatal activity scales across multiple intervals. Third, striatal ramping neurons are correlated with activity of the medial frontal cortex. Finally, interval timing and striatal ramping activity are disrupted when the medial frontal cortex is inactivated. Our results support the view that striatal neurons integrate medial frontal activity and are consistent with drift-diffusion models of interval timing. This principle elucidates temporal processing in frontostriatal circuits and provides insight into how the medial frontal cortex exerts top-down control of cognitive processing in the striatum. SIGNIFICANCE STATEMENT The ability to guide actions in time is essential to mammalian behavior from rodents to humans. The prefrontal cortex and striatum are critically involved in temporal processing and share extensive neuronal connections, yet it remains unclear how these structures represent time. We studied these two brain areas in rodents performing interval-timing tasks and found that time-dependent “ramping” activity, a monotonic increase or decrease in neuronal activity, was a key temporal signal. Furthermore, we found that striatal ramping activity was correlated with and dependent upon medial frontal activity. These results provide insight into information-processing principles in frontostriatal circuits.

Schizophrenia involves abnormalities in the medial frontal cortex that lead to cognitive deficits. Here we investigate a novel strategy to normalize medial frontal brain activity by stimulating cerebellar projections. We used an interval timing task to study elementary cognitive processing that requires both frontal and cerebellar networks that are disrupted in patients with schizophrenia. We report three novel findings. First, patients with schizophrenia had dysfunctional delta rhythms between 1–4 Hz in the medial frontal cortex. We explored cerebellar-frontal interactions in animal models and found that both frontal and cerebellar neurons were modulated during interval timing and had delta-frequency interactions. Finally, delta-frequency optogenetic stimulation of thalamic synaptic terminals of lateral cerebellar projection neurons rescued timing performance as well as medial frontal activity in a rodent model of schizophrenia-related frontal dysfunction. These data provide insight into how the cerebellum influences medial frontal networks and the role of the cerebellum in cognitive processing.