Publications using TDT

Effective regulation of energy metabolism is critical for survival. Metabolic control involves various nuclei within the hypothalamus, which receive information about the body’s energy state and coordinate appropriate responses to maintain homeostasis, such as thermogenesis, pancreatic insulin secretion, and food-seeking behaviors. It has recently been found that the hippocampus, a brain region traditionally associated with memory and spatial navigation, is also involved in metabolic regulation. Specifically, hippocampal sharp wave-ripples (SWRs), which are high-frequency neural oscillations supporting memory consolidation and foraging decisions, have been shown to reduce peripheral glucose levels. However, whether SWRs are enhanced by recent feeding—when the need for glucose metabolism increases, and if so, whether feeding-dependent modulation of SWRs is communicated to other brain regions involved in metabolic regulation—remains unknown. To address these gaps, we recorded SWRs from the dorsal CA1 region of the hippocampus of mice during sleep sessions before and after consumption of meals of varying caloric values. We found that SWRs occurring during sleep are significantly enhanced following food intake, with the magnitude of enhancement being dependent on the caloric content of the meal. This pattern occurred under both food-deprived and ad libitum feeding conditions. Moreover, we demonstrate that GABAergic neurons in the lateral hypothalamus, which are known to regulate food intake, exhibit a robust SWR-triggered increase in activity. These findings identify the satiety state as a factor modulating SWRs and suggest that hippocampal-lateral hypothalamic communication is a potential mechanism by which SWRs could modulate peripheral metabolism and food intake.

Gain-of-function (GoF) variants in the GRIN2D gene, encoding the GluN2D subunit of the N-methyl-D-aspartate receptor (NMDAR), cause a severe developmental and epileptic encephalopathy (DEE) characterized by intractable seizures, hypotonia and neurodevelopmental delay. We generated mice carrying the GoF V664I variant, orthologous to V667I, which is present in ∼25% of GRIN2D-DEE patients. Heterozygous mutant mice demonstrate behavioural, neuroanatomical and electrophysiological abnormalities. Lethal convulsive seizures are observed beginning at postnatal Day 17. As adults, heterozygotes display abundant and prolonged runs of spike-wave discharges (SWD) that often persist for minutes. The SWD epochs consist of different populations, differentiated by frequency and association with time-locked behavioural arrest. V664I mutant neurons have enlarged presynaptic terminals and increased synaptic distance. Functional analysis reveals increased inhibitory synaptic activity without changes in NMDAR decay kinetics or presynaptic plasticity in CA1 neurons, and analysis of hippocampal local field potentials shows a 1.5-fold increase in evoked responses and a 1.7-fold increase in action potential generation. Notably, expression of V664I in GABAergic interneurons, but not excitatory forebrain neurons, is sufficient to recapitulate the severe electroclinical phenotype. Altogether, our studies show that altered NMDAR function in inhibitory neurons plays a prominent role in DEE associated with GRIN2D GoF variants, suggesting that targeted genetic treatment may represent a path forward to successful therapeutic intervention.

Distinguishing between nectar and non-nectar odors is challenging for animals due to shared compounds and varying ratios in complex mixtures. Changes in nectar production throughout the day and over the animal’s lifetime add to the complexity. The honeybee olfactory system, containing fewer than 1000 principal neurons in the early olfactory relay, the antennal lobe (AL), must learn to associate diverse volatile blends with rewards. Previous studies identified plasticity in the AL circuits, but its role in odor learning remains poorly understood. Using a biophysical computational model, tuned by in vivo electrophysiological data, and live imaging of the honeybee’s AL, we explored the neural mechanisms of plasticity in the AL. Our findings revealed that when trained with a set of rewarded and unrewarded odors, the AL inhibitory network suppresses responses to shared chemical compounds while enhancing responses to distinct compounds. This results in improved pattern separation and a more concise neural code. Our calcium imaging data support these predictions. Analysis of a graph convolutional neural network performing an odor categorization task revealed a similar mechanism for contrast enhancement. Our study provides insights into how inhibitory plasticity in the early olfactory network reshapes the coding for efficient learning of complex odors.

In bats, which express a complex vocal repertoire and are considered vocal learners, the frontal auditory field (FAF) is supposedly placed in a frontal cortico‐striatal network for vocal–motor control. The FAF receives input from the auditory cortex (AC) and other auditory nuclei via multiple pathways. However, not much is known about the transition of information on vocalizations from the AC to the FAF. The bat AC consists of different subfields, among which the dorsal fields (dAC) are characterized by precise coding of the temporal envelope of vocalizations. The dAC should, therefore, be a major source of auditory feedback information about self‐produced or perceived vocalizations to the FAF. Our study aimed to investigate the specificity of encoding for different types of vocalizations in FAF and dAC neurons. Using extracellular recordings in anesthetized Phyllostomus discolor , we describe basic response properties in both cortical areas and compare responses to different types of prerecorded vocalizations. The specificity of encoding for different behaviorally relevant call categories and single calls was higher in dAC than in FAF neurons, both in terms of temporal firing patterns and response strength. These findings highlight the importance of the dAC in the neural network for control of vocal communication in bats.

While evolutionary cortical expansion is thought to underlie the evolution of human cognitive capabilities, excessive developmental expansion can lead to megalencephaly, often found in neurodevelopmental disorders. Still, little is known about how the overproduction of cortical neurons during development affects cortical processing and behavior in later life. Here we show that developmental overproduction of cortical superficial neurons impairs auditory processing in adult mice. We applied XAV939 to overproduce cortical superficial excitatory neurons during development. XAV939-treated adult mice exhibited auditory behavioral deficits and abnormal auditory cortical processing. Furthermore, we found fewer functional monosynaptic connections between cortical putative excitatory neurons. Altogether, our results suggest that abnormal auditory cortical processing contributes to the atypical auditory detectability in XAV939-treated mice. Although the expansion of cortical size is evolutionarily advantageous, an abnormal expansion during development can result in detrimental effects on cortical processing and perceptual behavior in adulthood.

Introduction Age-related hearing loss (ARHL) is linked to dementia, with mitochondrial dysfunction playing a key role in its progression. Deficient mitochondrial tRNA modifications impair protein synthesis and energy metabolism, accelerating ARHL. Mitochonic acid 5 (MA-5) has shown promise as a therapeutic candidate by improving mitochondrial function, reducing oxidative stress, and stabilizing membrane potential. Methods In this study, we investigated the effects of MA-5 on ARHL in cyclin-dependent kinase 5 regulatory subunit-associated protein 1 ( Cdk5rap1 ) knockout (KO) mice, which exhibit early-onset ARHL due to abnormalities in mitochondrial transfer RNA (mt-tRNA) modifications. Results MA-5 treatment effectively attenuated ARHL progression in Cdk5rap1 -KO mice by improving auditory brainstem response thresholds and distortion product otoacoustic emissions. It also reduced spiral ganglion and outer hair cell loss, while preserving the cochlear structural integrity by preventing mitochondrial degeneration in spiral ligament fibrocytes. Mechanistically, MA-5 upregulated the expression of silent information regulator sirtuin 1 and promoted the nuclear translocation of yes-associated protein, both of which are involved in regulating mitochondrial function and cellular senescence. Metabolomics analysis further demonstrated that MA-5 restored mitochondrial metabolism, reduced lactate accumulation, and maintained mitochondrial integrity. Conclusion These findings suggest that MA-5 is a viable treatment option for ARHL and other age-related disorders associated with mitochondrial dysfunction.

Auditory learning is supported by long-term changes in the neural processing of sound. We examined these task-depend changes in the auditory cortex by mapping neural sensitivity to timbre, pitch, and location cues in cues in trained ( n = 5) and untrained control female ferrets ( n = 5). Trained animals either identified vowels in a two-alternative forced choice task ( n = 3) or discriminated when a repeating vowel changed in identity or pitch ( n = 2). Neural responses were recorded under anesthesia in two primary auditory cortical fields and two tonotopically organized nonprimary fields. In trained animals, the overall sensitivity to sound timbre was reduced across three cortical fields compared with control animals, but maintained in a nonprimary field (the posterior pseudosylvian field). While training did not increase sensitivity to timbre across the auditory cortex, it did change the way in which neurons integrated spectral information, with neural responses in trained animals increasing their sensitivity to first and second formant frequencies, whereas in control animals cortical sensitivity to spectral timbre depended mostly on the second formant. Animals trained on timbre identification were required to generalize across pitch when discriminating timbre, and their neurons became less modulated by fundamental frequency relative to control animals. Finally, both trained groups showed increased spatial sensitivity and an enhanced response to sound source locations close to the midline, where the loudspeaker was located in the training chamber. These results demonstrate that training elicited widespread alterations in the cortical representation of complex sounds.

Introduction Ibrutinib, widely used in leukemia treatment, has been implicated in sensorineural hearing loss; however, its underlying mechanisms remain unclear. Methods This study investigated the impact of ibrutinib on hearing using HEI-OC1 cells, cochlear explants and C57BL/6 J mice. We used RNA-sequences analysis to investigate the potential mechanisms of ibrutinib-induced ototoxicity. Mice received ibrutinib and auditory thresholds were assessed via auditory brainstem response testing; to assess the potential protective effects, we co-administered the caspase inhibitor Z-Val-Ala-Asp (OMe)-fluoromethylketone (Z-VAD-FMK) and monitored hearing. Results Z-VAD-FMK mitigated ibrutinib-induced hearing loss by inhibiting apoptosis in auditory cells. Ibrutinib exposure resulted in cochlear hair cell (HC) damage and subsequent hearing loss by inhibiting the protein kinase B and G protein-coupled receptor 83 (GPR83) pathways. RNA sequencing suggested that GPR83 protects HCs by modulating autophagy. Z-VAD-FMK application and GPR83 overexpression attenuated ibrutinib-induced cochlear HC apoptosis and auditory decline. Conclusion These findings confirm ibrutinib’s ototoxicity and highlight the protective role of GPR83 in ibrutinib-induced hearing loss, supporting future clinical investigations into Z-VAD-FMK and GPR83 as interventions for ibrutinib or other chemotherapeutic drug-induced ototoxicity.

Perception of rhythm significantly impacts various aspects of daily life, including engaging with music, discerning speech prosody nuances, and coordinating physical activities like walking and sports. Numerous studies in cognitive sciences have highlighted that human rhythmic synchronization is more precise when responding to auditory rhythmic stimuli than to visual ones when the timing cues are identical. However, deaf individuals were shown to display a heightened proficiency in synchronizing their movements with visual timing cues, outperforming hearing controls (HC). Furthermore, it was demonstrated that cochlear implant (CI) users can synchronize their movements with the rhythm of unpitched drum tones. These findings raise an important question: do CI users possess a visual synchronization advantage from their pre-implant deafness, while maintaining auditory synchronization skills comparable to those of HC? Alternatively, does the neural reorganization post-implantation negate the visual synchronization advantage acquired before the implant? This study aims to answer these questions by using a sensorimotor synchronization task to probe multisensory processing abilities in CI users. Specifically, we assessed unimodal and multimodal auditory and visual abilities in CI users compared to HC using a finger tapping synchrony task with four isochronous stimulus conditions: an auditory metronome, a visual metronome, a synchronous presentation of both the auditory and visual metronomes at the same tempo, and an asynchronous presentation of the auditory and visual stimuli at differing tempos. Synchronization to auditory stimuli surpassed synchronization to visual stimuli in both groups. CI users and HC demonstrated similar unisensory synchronization consistency within the visual and auditory conditions. While HC enhanced their consistency in the audio-visual synchronous condition compared to the unisensory visual condition, CI users did not display the same improvement. Furthermore, the interference from incongruent auditory information in the asynchronous condition was comparable in HC and CI users. This study highlights that, although pitch processing is known to be impaired in CI users, our findings suggest that rhythm processing remains relatively spared. As anticipated, CI users demonstrate similar auditory rhythmic synchronization skills to those of HC, in line with existing research. Moreover, we find that, unlike deaf individuals, CI users do not exhibit an advantage in visual rhythmic synchronization, which may be due to the relatively few CI users in the study who had early prolonged pre-implantation deafness. The observed shift in audio-visual integration among CI users suggests that post-deafness or post-implantation reorganization of their auditory cortex may impede the effective integration of temporal auditory stimulation from the implant and visual information.

Exposure to traumatic noise triggers cochlear damage and consequently causes permanent sensorineural hearing loss. However, effective treatment strategies for noise-induced hearing loss (NIHL) are lacking. Sirtuin 1 (SIRT1) is a NAD+-dependent deacetylase that plays a critical role in multiple physiological and pathological events. However, its role in NIHL pathogenesis remains elusive. This study revealed that SIRT1 expression in the cochlea progressively decreases in a mouse model of NIHL. Hair cell-specific knockout of SIRT1 exacerbates the noise-induced loss of outer and inner hair cell synaptic ribbons, retraction of cochlear nerve terminals, and oxidative stress, leading to more severe NIHL. Conversely, adeno-associated virus (AAV)-mediated SIRT1 overexpression effectively attenuated most noise-induced cochlear damage and alleviated NIHL. Transcriptomic analysis revealed that SIRT1 deficiency impairs glucose metabolism and inhibits antioxidant pathways in the cochlea following exposure to noise. Further investigation revealed that SIRT1 exerts an antioxidant effect, at least in part, through AMPK activation in cultured auditory HEI-OC1 cells exposed to oxidative stress. Collectively, these findings indicate that SIRT1 is essential for the maintenance of redox balance and mitochondrial function in the cochlea after traumatic noise exposure, thus providing a promising therapeutic target for NIHL treatment.

The mesolimbic dopamine system is a set of subcortical brain circuits that plays a key role in reward processing, reinforcement, associative learning, and behavioral responses to salient environmental events. In our previous studies of the dopaminergic response to salient visual stimuli, we observed that dopamine release in the lateral nucleus accumbens (LNAc) of mice encoded information about the rate and magnitude of rapid environmental luminance changes from darkness. Light-evoked dopamine responses were rate-dependent, robust to the time of testing or stimulus novelty, and required phototransduction by rod and cone opsins. However, it is unknown if these dopaminergic responses also involve non-visual opsins, such as melanopsin, the primary photopigment expressed by intrinsically photosensitive retinal ganglion cells (ipRGCs). In the current study, we evaluated the role of melanopsin in the dopaminergic response to light in the LNAc using the genetically encoded dopamine sensor dLight1 and fiber photometry. By measuring light-evoked dopamine responses across a broad irradiance and wavelength range in constitutive melanopsin ( Opn4 ) knockout mice, we were able to provide new insights into the ability of non-visual opsins to regulate the mesolimbic dopamine response to visual stimuli.

Animals learn the value of foods on the basis of their postingestive effects and thereby develop aversions to foods that are toxic 1–10 and preferences to those that are nutritious 11–13 . However, it remains unclear how the brain is able to assign credit to flavours experienced during a meal with postingestive feedback signals that can arise after a substantial delay. Here we reveal an unexpected role for the postingestive reactivation of neural flavour representations in this temporal credit-assignment process. To begin, we leverage the fact that mice learn to associate novel 14,15 , but not familiar, flavours with delayed gastrointestinal malaise signals to investigate how the brain represents flavours that support aversive postingestive learning. Analyses of brain-wide activation patterns reveal that a network of amygdala regions is unique in being preferentially activated by novel flavours across every stage of learning (consumption, delayed malaise and memory retrieval). By combining high-density recordings in the amygdala with optogenetic stimulation of malaise-coding hindbrain neurons, we show that delayed malaise signals selectively reactivate flavour representations in the amygdala from a recent meal. The degree of malaise-driven reactivation of individual neurons predicts the strengthening of flavour responses upon memory retrieval, which in turn leads to stabilization of the population-level representation of the recently consumed flavour. By contrast, flavour representations in the amygdala degrade in the absence of unexpected postingestive consequences. Thus, we demonstrate that postingestive reactivation and plasticity of neural flavour representations may support learning from delayed feedback.

Episodic memory requires the precise coordination between the hippocampus and distributed cortical regions. This may be facilitated by bursts of brain activity called high-frequency oscillations (HFOs). We hypothesized that HFOs activate specific networks during memory retrieval and aimed to describe the electrophysiological properties of HFO-associated activity. To study this, we recorded intracranial electroencephalography while human participants performed a list learning task. Hippocampal HFOs (hHFOs) increased during encoding and retrieval, and these increases correlated with memory performance. During retrieval, hHFOs demonstrated activation of semantic processing regions that were previously active during encoding. This consisted of broadband high-frequency activity (HFA) and cortical HFOs. HFOs in the anterior temporal lobe, a major semantic hub, co-occurred with hHFOs, particularly during retrieval. These coincident HFOs were associated with greater cortical HFA and cortical theta bursts. Hence, HFOs may support synchronization of activity across distributed nodes of the hippocampal-cortical memory network.

Transmembrane protein 135 (TMEM135) is a highly conserved 52 kDa protein with five predicted transmembrane domains that colocalizes with mitochondria and peroxisomes. Previous studies have shown that TMEM135 is involved in mitochondrial dynamics, thermogenesis, and lipid metabolism across multiple tissues and species; however, its role in the inner ear and auditory system remains unknown. We investigated the function of TMEM135 in hearing using wild-type (WT) and Tmem135FUN025/FUN025 (FUN025) mutant mice on a CBA/CaJ background, a normal-hearing mouse strain. Although FUN025 mice displayed normal auditory brainstem response (ABR) thresholds at 1 month, we observed significantly elevated ABR thresholds at 8, 16, and 64 kHz by 3 months, which progressed to profound hearing loss by 12 months. Consistent with our auditory testing results, 13-month-old FUN025 mice exhibited a severe loss of outer hair cells and more modest changes in inner hair cell survival, spiral ganglion neuron density, and stria vascularis integrity in the cochlea. Our results using BaseScope RNA in situ hybridization indicate that TMEM135 is expressed in the inner hair cells, outer hair cells, supporting cells, and stria vascularis. Using Volocity software and Costes colocalization analysis, we found that TMEM135 closely colocalizes with mitochondria in hair cells. Together, these results demonstrate that the FUN025 mutation in Tmem135 causes progressive sensorineural hearing loss, and suggest that TMEM135 is crucial for maintaining key cochlear cell types and normal sensory function in the aging cochlea.

Odorants play a critical role in regulating feeding behavior by signaling potential threats or food sources in the environment. However, the neural mechanisms by which odorants affect feeding are not well understood. Tachykinin-1-expressing neurons in the parasubthalamic nucleus (PSTNTac1 neurons) are critical for reducing food intake in response to internal appetite-suppressing hormones, gastric distension, and external cues that signal danger. Therefore, we tested the hypothesis that activity in these neurons is modulated by exposure to aversive, attractive, and neutral odorants. Using fiber photometry in mice, we found that PSTNTac1 neurons increase activity in response to the aversive predator odorants 2-methyl-2-thiazoline (2MT) and 2,5-dihydro-2,4,5-trimethylthiazoline (TMT), but not to neutral or attractive odorants. This activation correlates with a reduction in food intake and an increase in the latency to initiate feeding. Furthermore, chemogenetic inhibition of PSTNTac1 neurons blocks the suppression of feeding caused by 2MT and TMT. These findings highlight the specificity of PSTNTac1 neurons in processing aversive olfactory signals and their critical role in integrating external threat cues with internal signals that regulate appetite.

The cochlea is vulnerable to various pathological conditions, with sensory cells typically being the primary targets of damage. However, supporting cells also experience significant impacts. Despite their critical role in maintaining the structural and functional integrity of the sensory epithelium, the supporting cell involvement in cochlear damage remains poorly understood. This study aimed to elucidate the susceptibility of supporting cells in cochlear damage and their role in structural repair, using a mouse model of ototoxicity induced by cyclodextrin-a cyclic oligomer of glucose that is known to preferentially damage outer hair cells at high doses. A morphological examination of the cochlea showed that cyclodextrin exposure caused significant sensory cell loss, particularly affecting outer hair cells across the cochlear spiral, except at the apex. Despite extensive hair cell damage, most supporting cells in the apical and middle cochlear regions survived. In the basal end, where substantial supporting cell loss occurred, certain Deiters' cells survived even after losing their phalangeal processes. Additionally, our observations indicate that Hensen's cells contribute to forming an epithelial layer over the basilar membrane when the organ of Corti collapses. Further quantitative analysis revealed location-dependent susceptibility among supporting cell types. Deiters' cells demonstrated greater resilience than pillar cells. Notably, the three rows of Deiters' cells displayed differential susceptibility: the third row showed a more significant loss in regions with sporadic Deiters' cell loss, while the first row exhibited an increased loss in areas adjacent to regions of complete Deiters' cell depletion. The reduction of Hensen's cells started in the middle section of the cochlea, occurring at a greater level than the reduction observed in Deiters' and pillar cells. However, in the extreme base, where both pillar and Deiters' cells were largely or completely absent, some Hensen's cells were still present. Together, these findings provide new insights into the varying vulnerability of supporting cells to cochlear damage and underscore their essential role in structural repair.

A failure of organized communication in the PFC is thought to contribute to the emergence of cognitive impairments in psychiatric diseases, with attentional deficits occurring as a fundamental symptom across various conditions. The 22q11.2 microdeletion syndrome is a rare genetic condition that confers a high risk for developing psychiatric and neurodevelopmental disorders, and mouse models have been shown to display attention impairments and PFC pathology that are relevant to clinical populations. Abnormalities in prefrontal parvalbumin-expressing neurons (PVNs) are part of the observed pathophysiology, and studies in rodents have shown that the direct manipulation of these cells can induce behavioral deficits that align with the cognitive symptoms observed in psychiatric diseases. In the present study, we expanded on the role of PVNs in supporting cognition by investigating their involvement in multiple aspects of attentional functions using a translationally relevant task of focused visual attention, in both non-pathological mice and a model of the 22q11.2 microdeletion syndrome. We observed that task-evoked prefrontal PVN activity was reduced in mice that exhibited poorer attention and in 22q11.2 mutant mice. While PVN activity was shaped across learning in non-pathological mice, mutant mice exhibited a lack of signal dynamics that coincided with attentional deficits. Importantly, we observed that task performance in both poor performing wild-types and 22q11.2 mutants could be alleviated by gamma frequency stimulation of PVNs. Thus, PVNs appear to be involved in the acquisition of task rules and execution of attention and continue to be a promising therapeutic target for cognitive dysfunction in disease.

Early and accurate diagnosis of Alzheimer's disease (AD) will be key for effective personalized treatment plans ( Cummings, 2023). Significant difficulties in auditory processing have been frequently reported in many patients with mild cognitive impairment, the prodromal form of AD ( Tarawneh et al., 2022), making it an outstanding candidate as AD diagnostic biomarker. However, the efficiency of diagnosis with this parameter has not been explored. Here we show that when male mice with amyloidosis begin to show memory decline, changes in the auditory brainstem response (ABR) to clicks enable the reliable diagnosis of disease using a machine learning algorithm. Interpretation of the machine learning diagnosis revealed that the upper levels of the auditory pathway, including the inferior colliculus, were the probable sources of the defects. Histological analyses show that in these locations, neuroinflammation and plaque deposition temporally correlate with behavioral changes consistent with memory loss. While these findings are tempered by the caveat that they derive from amyloidosis mice, we propose that ABR measurements be evaluated as an additional rapid, low-cost, noninvasive biomarker to assist the diagnostic testing of early-stage AD.

Aims This study examined the effect of 4,4′‐diisothiocyanostilbene‐2,2′‐disulfonic acid (DIDS), an anion channel blocker of voltage‐dependent anion channel 1 (VDAC1), on noise‐induced hearing loss (NIHL) and its underlying mechanisms. Methods Cochlear explants and House Ear Institute‐Organ of Corti 1 (HEI‐OC1) cells were used to assess the effect of DIDS in vitro. Auditory brainstem responses were used to assess auditory functions in mice. Immunofluorescence staining of myosin 7a and CTBP2 were used to examine hair cells and synaptic ribbons. The accumulation of reactive oxygen species (ROS) was measured by 4‐HNE staining. The gene expression changes of cochlea were analyzed using RNA sequencing. Results DIDS reduced the levels of ROS in cochlear explants and attenuated cell death caused by hydrogen peroxide in both cochlear explants and HEI‐OC1 cells. In C57BL/6 mice, DIDS reduced ROS generation and tumor necrosis factor‐α induced by noise exposure, thereby protecting outer hair cells and inner hair cell synaptic ribbons from noise‐induced damage through a mechanism involving the PINK1/Parkin signaling pathway. The preventive effect of DIDS in cochlear explants was eliminated by mitophagy inhibition. Conclusion VDAC1 inhibition enhances mitophagy in cochlear hair cells, playing a critical role in defending against oxidative stress and inflammation. Downregulation of VDAC1 may thus be considered a therapeutic strategy for preventing cochlear hair cell damage and reducing NIHL.

Dopamine (DA) and serotonin (5-HT) are neuromodulators in reward processing, decision- making, and motivated behavior. While often viewed as opposing or complementary systems, how DA and 5-HT release integrate in the striatum remains elusive. Using optogenetics, fiber photometry, and slice electrophysiology, we found that ventral tegmental area (VTA) DA neuron stimulation increased DA release without affecting 5-HT release. Dorsal raphe nucleus (DRN) 5-HT neuron activation, on the other hand, induced serotonin release and a transient increase in DA in the NAc, likely via glutamate co-release onto VTA DA neurons during the initial stimulation phase. These findings indicate that DA and 5-HT operate largely independently in the striatum, with selective circuit-dependent interactions. This work refines our understanding of DA-5HT interactions and provides a foundation for future research into their roles in motivated behaviors and neuropsychiatric disorders.