Functional autapses form in striatal parvalbumin 3 interneurons but not medium spiny neurons

21 Autapses (or self-synapses) selectively form in specific cell types in many brain regions including the neocortex and the hippocampus, where they provide feedback control 23 over self-spiking activities. Previous morphological studies also found putative 24 autapses in medium spiny neurons (MSNs) of the striatum. However, it remains unclear 25 whether striatal neurons indeed form physiologically functional autapses. We 26 performed whole-cell recordings from striatal neurons in acute mouse brain slices, and 27 identify autaptic neurons by the occurrence of prolonged asynchronous release (AR) of 28 neurotransmitter after high-frequency burst of action potentials (APs) in the same cell. 29 To our surprise, we found no autaptic release in all recorded MSNs after the AP burst, 30 even in the presence of Sr 2+ that should desynchronize and thus prolong synaptic vesicle 31 release. In sharp contrast, we observed robust autaptic AR events in half of the recorded 32 parvalbumin (PV)-positive neurons. Autaptic responses in PV cells were mediated by 33 GABA A receptors, and the AR strength was dependent on the frequency and the number 34 of APs during the burst. Further simulation results show that autapses regulate burst spiking in PV cells by providing self-inhibition and thus shape network oscillation at 36 certain frequencies. Together, we reveal that, distinct from MSNs, striatal PV neurons 37 form functional autapses, activation of which would regulate self-activities in PV cells, 38 and thereby shape MSN firing and network oscillations. 42 communication in the nervous system. However, some types of neurons form autapses, 43 where a neuron synapses onto itself. Autaptic transmission provides feedback signal 44 regulating self-spiking activities. Neuronal and network activities in the striatum play 45 critical roles in motor control and other brain functions. Previous studies suggest 46 formation of autapses in striatal principal MSNs, but it remains unclear whether striatal 47 neurons form functional autapses. We performed direct recordings from striatal neurons 48 and examined the occurrence of autaptic transmission in acute brain slices. Surprisingly, 49 we did not detect any autaptic responses in MSNs. A large proportion of striatal PV 50 neurons, however, produced robust autaptic GABA release upon high-frequency 51 stimulation, indicating selective formation of autapses in striatal PV cells. Our 52 computation simulations suggest that autapses provide self-inhibition in PV cells and 53 thereby shape activities in MSNs and striatal network, particularly when PV cells 54 discharge at high frequencies corresponding to a high dopamine state. Together, our 55 findings indicate that PV cells, but not MSNs, in the striatum form physiologically 56 functional autapses. Autapses in PV cells could be essential circuit elements in the striatum and contribute to striatal functions, such as motor control.


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Striatum is the largest nucleus in the basal ganglia receiving synaptic inputs from different cortical areas, thalamic nuclei and limbic regions [1]. It  Autapses form in striatal PV cells 156 We performed similar experiments in PV cells with tdTomato expression. Close 157 examination of their morphology revealed that PV cells possessed smooth dendrites 158 and dense axon collaterals (Fig 2A and B), similar to previous studies [7,9,28]. PV  from CCK cells than that from PV neurons [30]. To exclude the possibility that some 215 autaptic PV cells might have only SR, but no detectable AR, we added SrCl 2 (5 mM, 216 see Methods) to the bath solution so that autaptic GABA release could be 217 desynchronized [21] (Fig 3D). In our experiments, the presence of Sr 2+ significantly increased the PT-AR duration from 387 ± 43 to 586 ± 63 ms (control, n = 27; Sr 2+ , n = 219 21, P = 9.97×10 -3 , two-sample Student's t test). The number of AR events also slightly 220 increased from 13.1 ± 1.7 to 15.8 ± 2.1, but with no significant difference (P = 0.31,

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Wilcoxon rank sum test, Fig 3E). 222 We next sought to examine the percentage of PV cells that form autapses. In 223 recorded PV cells, we applied high-frequency stimulations (20- to that in control condition (without Sr 2+ , 46.6%, n = 61/131, Fig 3F), and also similar 228 to that found in cortical PV neurons [19]. These probabilities should be underestimated 229 because slice preparation reduced the complexity of neuronal dendrites and axons.         adding AR showed no further effect on these spiking properties. At the high DA level 303 (Fig 5), we found that SR alone reduced the average firing rate (n = 10 trials, P = 5.75 304 ×10 -5 , two-sample Student's t test) and the average duration of bursts (P = 2.00×10 -3 , 305 two-sample Student's t test), but had no significant effect on the burst interval (P = 0.47, Wilcoxon rank sum test, Fig 5G). When we added AR to the autapse (SR+AR), we also 307 observed a slight decrease in the firing rate (n = 40 trials, P = 1.21×10 -9 , ANOVA) and 308 the burst duration (P = 1.31×10 -3 ) as the AR strength increased from 0 to 2. Increasing 309 AR strength also prolonged the interval between bursts significantly (P = 7.87×10 -3 , 310 Fig 5H). significantly increased (Fig 6E and G). In following simulations, we added autaptic AR to PV cells. We found that autaptic 353 AR slightly decreased the firing rate of PV neurons (n = 40, P = 3.18×10 -5 , ANOVA, 354 Fig 6H). In sharp contrast to SR alone, AR had no effect on burst duration (P = 0.563, 355 ANOVA) but slightly increased the burst interval (P = 6.61×10 -5 , ANOVA, Fig 6H). A  In this study, we show that autaptic contacts occur in PV interneurons in the striatum.  added SR. However, in the presence of AR, both theta and gamma power were significantly increased (Fig 6). Since AR only occur at high frequencies of PV cells, it 417 should only play a role at high DA levels. As expected, adding AR showed no additional 418 effect on neuronal and network activity when PV cells discharge at low frequencies (i.e.

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baseline DA level). Therefore, we mainly focused on physiological contribution of 420 autapses at states when PV cell discharge at high frequencies. Since PV cells show fast-spiking firing pattern and are able to discharge up to 220 Hz 426 (Fig 2), they contribute largely to high-frequency LFP oscillations. Indeed, we found a 427 decrease in PV cell spike frequency and a shift of power density from high gamma to 428 other bands after adding autaptic SR (Fig 6E-G). Since the frequency of AR events  Table 1. The steady-state of gating variable The time constant of decay = 1/( + )

PV cell (two compartments)
The  . To examine the contribution of autapses to network oscillation, we normalized (z-527 score) the simulated LFP before plotting the power spectrum. All simulations were run 528 on MATLAB software (version R2021a). All differential equations were integrated 529 using a fourth-order Runge-Kutta algorithm with time step 0.01 ms.  For two independent observations with normal distribution (P > 0.05, Shapiro Tukey's test for post-hoc analysis, were used for comparisons of multiple groups.

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Datasets were considered to be significantly different if P < 0.05.