The nanoscopic organization and regulation of individual molecular components in presynaptic varicosities of neurons releasing modulatory volume neurotransmitters like dopamine (DA) remain largely elusive. Here we show by application of several single-molecule sensitive super-resolution microscopy techniques to cultured neurons and mouse striatal slices, that the dopamine transporter (DAT), a key protein in varicosities of dopaminergic neurons, exists in the membrane in dynamic equilibrium between an inward-facing nanodomain-localized and outward-facing unclustered configuration. The balance between these configurations is inversely regulated by excitatory drive and by DA D2-autoreceptor activation in manner dependent on Ca2+-influx via N-type voltage-gated Ca2+-channels. The DAT nanodomains contain tens of transporters molecules and overlap with nanodomains of PIP2 (phosphatidylinositol-4,5-bisphosphate) but show little overlap with D2-autoreceptor, syntaxin-1 and clathrin nanodomains. By demonstrating that nanoscopic reorganizations with putative major impact on transmitter homeostasis can take place in dopaminergic varicosities, the data have important implications for understanding modulatory neurotransmitter physiology.
The dorsal (DS) and ventral striatum (VS) receive dopaminergic projections that control motor functions and reward-related behavior. It remains poorly understood how dopamine release dynamics across different temporal scales in these regions are coupled to behavioral outcomes. Here, we employ the dopamine sensor dLight1.3b together with multiregion fiber photometry and machine learning-based analysis to decode dopamine dynamics across the striatum during self-paced exploratory behavior in mice. Our data show a striking coordination of rapidly fluctuating signal in the DS, carrying information across dopamine levels, with a slower signal in the VS, consisting mainly of slow-paced transients. Importantly, these release dynamics correlated with discrete behavioral motifs, such as turns, running, and grooming on a subsecond-to-minute time scale. Disruption of dopamine dynamics with cocaine caused randomization of action selection sequencing and disturbance of DS–VS coordination. The data suggest that distinct dopamine dynamics of DS and VS jointly encode behavioral sequences during unconstrained activity with DS modulating the stringing together of actions and VS the signal to initiate and sustain the selected action.
A fundamental concept in neuroscience is the transmission
of information
between neurons via neurotransmitters, -modulators, and -peptides.
For the past decades, the gold standard for measuring neurochemicals
in awake animals has been microdialysis (MD). The emergence of genetically
encoded fluorescence-based biosensors, as well as in vivo optical techniques such as fiber photometry (FP), has introduced
technologically distinct means of measuring neurotransmission. To
directly compare MD and FP, we performed concurrent within-animal
recordings of extracellular dopamine (DA) in the dorsal striatum (DS)
before and after administration of amphetamine in awake, freely behaving
mice expressing the dopamine sensor dLight1.3b. We show that despite
temporal differences, MD- and FP-based readouts of DA correlate well
within mice. Down-sampling of FP data showed temporal correlation
to MD data, with less variance observed using FP. We also present
evidence that DA fluctuations periodically reach low levels, and naïve
animals have rapid, predrug DA dynamics measured with FP that correlate
to the subsequent pharmacodynamics of amphetamine as measured with
MD and FP.
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