SK and Kv4 Channels Limit Spike Timing Perturbations in Pacemaking Dopamine Neurons

Abstract

Midbrain dopamine (DA) neurons are among the best characterized pacemaker neurons, having intrinsic, rhythmic firing activity even in the absence of synaptic input. However, the mechanisms of DA neuron pacemaking have not been systematically related to how these cells respond to synaptic input. The input-output properties of pacemaking neurons can be characterized by the phase resetting curve (PRC), which describes the sensitivity of inter-spike interval (ISI) length to inputs arriving at different phases of the firing cycle. Here we determined PRCs of putative DA neurons in the substantia nigra pars compacta (SNc) in brain slices from male and female mice using gramicidin-perforated current-clamp recordings with electrical noise stimuli applied through the patch pipette. On average, and compared to nearby putative GABA neurons, DA neurons showed a low, nearly constant level of sensitivity across most of the ISI, but individual cells had PRCs showing relatively greater sensitivity at early or late phases. Pharmacological experiments showed that DA neuron PRCs are shaped by small-conductance calcium-activated potassium (SK) and Kv4 channels, which limit input-sensitivity across early and late phases of the ISI. Our results establish the PRC as a tractable experimental measurement of individual DA neuron input-output relationships and identify two of the major ionic conductances that limit perturbations to rhythmic firing. These findings have applications in modeling and for identifying biophysical changes in response to disease or environmental manipulations.SIGNIFICANCE STATEMENTIn substantia nigra pars compacta dopamine neurons, pacemaking mechanisms determine the response to an instantaneous synaptic input according to the phase resetting curve (PRC). Here we measured PRCs of dopamine neurons and determined how they are shaped by small-conductance calcium-activated potassium (SK) and Kv4 channels, which regulate pacemaking rate and regularity. We found that both types of channel limit sensitivity to perturbations in firing. Thus, the currents responsible for slow pacemaking also control spike time responses to synaptic input.

VTA dopamine neurons are hyperexcitable in 3xTg-AD mice due to casein kinase 2-dependent SK channel dysfunction

“…NS309 reduced firing rate in WT, but not 3xTg neurons (Figure 4c). However, it had no significant impact on the CV of the ISI in either genotype (Figure 4d), contrary to our recent findings in SNc neurons 41 . When we further scrutinized ISI dynamics, we observed that WT neurons (Figure 4e) were more sensitive than 3xTg (Figure 4f) neurons; NS309 hyperpolarized the average mAHP (Figure 4g) and the minimum ISI voltage (Figure 4h) in WT but not 3xTg VTA DA neurons.…”

“…The mAHP is governed largely by action potential mediated influx of calcium and subsequent activation of SK channels 40 . Indeed, the reduced mAHPs and minimum ISI voltages in 3xTg mice were highly reminiscent of data we recently described in the SNc following a partial block of SK channels using apamin 41 , and a depolarized threshold may be secondarily linked to SK channels through sodium channel inactivation 38,42 . Thus, we aimed to determine whether SK channel activity is altered in 3xTg VTA DA neurons.…”

“…SK channel activation regulates both spontaneous firing and burst generation in midbrain DA neurons 14,30,80,81 . During tonic activity, SK channel activation during the ISI sets the timing for the next action potential and dampens sensitivity to synaptic input 41 . SK channel influence on motivated behavior may hinge on their physical proximity to NMDA receptors, where they can serve as a calcium-dependent shunt during excitatory synaptic input 80,81 .…”

“…Enhancing SK function normalizes firing of 3xTg DA neurons Positive allosteric modulators of SK channels have previously been shown to safeguard neurons from neurotoxic insults in degenerative models 48,49 . We next tested the sensitivity of VTA DA neurons by applying the selective SK channel positive allosteric modulator NS309 (1 µM) in perforated-patch recordings of WT (Figure 4a) and 3xTg VTA DA neurons (Figure 4b) 41,50 . NS309 reduced firing rate in WT, but not 3xTg neurons (Figure 4c).…”

Optogenetic action potentials and intrinsic pacemaker interplay in retrogradely identified midbrain dopamine neurons

“…In addition, potential effects of oAPs on intrinsic pacemaking have also not yet been studied in DA neurons. A systematic characterization of excitability changes across the pacemaker duty cycle of DA SN neurons in vitro has been recently probed by phase setting curves (Higgs et al, 2023). In this elegant study, the role of intrinsic conductances mediated by, for example, Kv4 and SK channels in limiting spike timing pertubations have been identified.…”