The specific function of a given neural circuit depends on the precise pattern of excitatory, inhibitory and neuromodulatory inputs on neuronal dendrites. The experiments presented in this study revealed that GABAergic interneurons use a microtubule-dependent strategy for synaptic development and organization of excitatory inputs. Immature interneuron dendrites generate protrusions as a means of searching the surrounding tissue microenvironment for appropriate axonal contacts (). After establishing nascent synaptic contacts, these protrusions serve as conduits for the retrograde synapse movement to the parental dendritic shafts (). Tight interaction of presynaptic and postsynaptic plasma membranes may transduce the tension generated in the dendritic protrusions to the presynaptic structure26
. After reaching the parental dendritic shafts, synapse translocation is downregulated (). Mixed polarity of microtubules in the parental dendritic shafts27
may suppress directional mobility of synapses. PSD maturation enhances NMDA receptor-dependent calcium influx28
, which may also negatively regulates PSD mobility. We also provided evidences indicating that precise regulation of dynein function by LIS1 and NDEL1 (ref. 21
) is critical for regulating synaptic mobility.
Model for coordinated remodelling of dendritic protrusions and PSDs in interneuron dendrites.
Various neuronal types in the brain develop dendrites without spines, but it is not clear whether transfer of synapses to dendritic shafts via specialized protrusions is a common mechanism. The aspiny dendrites of tectal neurons in zebrafish show a distinct pattern of PSD formation and dendrite development,29
in which new PSD puncta are formed exclusively on dendritic filopodia. Subsequent transformation of filopodia into dendritic branches is the major mechanism for synaptic development. The complex dendritic arbourization of zebrafish tectal neurons may be sufficient to maximize connectivity with surrounding axons. In contrast, the dendrites of mammalian cortical interneurons develop fewer branches, and their ability to search surrounding axons and form synapses would likely be limited if they did not extend long protrusions and then transfer the synaptic contacts to the dendritic shafts.
A more fundamental question is how dendritic protrusive activity and synapse maturation are controlled in specific neuronal types30
. The filopodia on pyramidal neurons may also function as conduits for retrograde synapse translocation, as indicated by imaging studies of retrograde PSD movement along such dendritic protrusions31
. Electron microscopic studies of pyramidal neurons in vivo
proposed the transfer of synapses formed onto filopodia-like protrusions to dendritic shafts7
. The different outcomes of synapse transfer between two neuron types may reflect the affinity of PSDs with cytoskeletal components. For example, pyramidal neuron PSDs may have a higher affinity for actin filaments, which could promote the formation of dendritic spines at the vicinity of PSDs after their transfer to the shafts.
We present several lines of evidence indicating that the dynamic behaviour of PSD puncta in interneuron dendrites is distinct from the mobility of non-synaptic scaffold complexes before their recruitment to synaptic contact sites. First, we observed the velocity of PSD puncta to be 2–5 μm h−1
, much slower than the reported velocity of non-synaptic scaffold complexes34
. Second, the direction of PSD translocation in interneurons was always towards the proximal ends of dendritic protrusions. In contrast, non-synaptic scaffold complexes reportedly move bidirectionally34
. Third, 78–91% of PSD puncta in interneuron dendritic protrusions also stained positively for presynaptic markers, indicating that most of the imaged PSD puncta were part of synaptic structures. Finally, our time-lapse imaging of presynaptic and postsynaptic structures using PSD-95-YFP and synaptophysin-CFP clearly demonstrated their coordinated movement. Collectively, these findings indicate that the motile behaviour of synaptic junctions on interneuron dendritic protrusions is distinct from the previously reported mobility of non-synaptic scaffold complexes.
The movement of PSDs along dendritic protrusions (2–5 μm h−1
) is slower than dynein-driven fast organelle transport35
, but comparable to slow high-load transport, such as nucleokinesis and centrosomal movement (5–10 μm h−1
. Both LIS1 and NDEL1 are known to be involved in dynein-dependent slow transport of intracellular structures21
, but their roles in rapid organelle transport have been reported to be minimal38
. LIS1 dysfunction leads to impairment of cell migration and nucleokinesis15
, reorientation of the entire microtubule system during mitosis41
, and aberrant centrosome and kinetochore dynamics42
. These previous data are consistent with the idea that the PSD translocation we reported here belongs to slow high-load transport and depends on LIS1 and NDEL1. LIS1 and NDEL1 may help dynein-dependent tethering of microtubules at the PSDs. Alternatively, LIS1 and NDEL1 may increase total force of multiple dynein molecules associated with PSDs21
. This mechanism may be required to overcome large mechanical resistance generated by interaction of PSDs with both presynaptic components and the surrounding cortical actin meshwork. Indeed, actin depolymerization using latrunculin A tended to enhance PSD movement along dendritic protrusions. Overall, our results indicate the essential role of postsynaptic dynein–NDEL1–LIS1 complex in slow high-load retrograde translocation of the entire synaptic junctions.
Our observations indicate that regulation of dynein function by the coordinated activity of LIS1 and NDEL1 (ref. 45
) is critical for determining the synaptic distribution on interneuron dendrites. Dysfunction of glutamatergic synapses on interneurons may lead to pathogenic disruption of the balance between excitatory and inhibitory synaptic transmission46
. Indeed, epilepsy is common in human type 1 lissencephaly, a genetic disorder caused by heterozygous loss or mutation of the Lis1
. We speculate that defects in LIS1-dependent regulation of synaptic mobility may promote epilepsy by disrupting excitatory inputs onto GABAergic interneurons in addition to the known effects of LIS-1 loss-of-function on the migration of early postmitotic neurons. This new aspect of LIS1 function may be important to our understanding of the pathophysiology of lissencephaly and help define new therapeutic targets.