Neuronal Cell Biology & Synaptic Function

The presynaptic compartment

From sensory perception to learning and memory, the functioning of the nervous system is dependent upon communication between neurons. Dysfunction of neuronal signaling results in neurological and neurodegenerative disorders ranging from autism to epilepsy and Alzheimer's disease. During neurotransmission at synapses specialized 40 nm secretory organelles termed synaptic vesicles undergo calcium-regulated fusion at the presynaptic active zone to release neurotransmitters that are recognized by neurotransmitter receptors at the postsynaptic side.   

The Haucke lab aims to address three fundamental questions regarding the formation and maintenance of the presynaptic compartment:

1) How are key presynaptic components such as synaptic vesicles and active zone proteins formed in the neuronal soma, axonally transported, and assembled into nascent synapses?
2) How and how fast are synaptic vesicle membranes internalized post-fusion and how are synaptic vesicles then reformed to maintain neurotransmission?
3) How is a functional synapse adapted and maintained in the mature and aging nervous system to sustain and plastically modulate neurotransmission, e.g. via neuronal autophagy or the endolysosomal system?
To address these questions, we combine optical imaging, suprer-resolution light and electron microscopy, live imaging, electrophysiology, and genetic approaches in primary mouse neurons and slice preparations and in human induced pluripotent stem cell derived neurons as models.

Formation of the presynaptic compartment

Our ability to move, to process sensory information or to form, store and retrieve memories crucially depends on the function of neuronal synapses. Synapses comprise a presynaptic compartment harboring the machinery for neurotransmitter release and an associated postsynaptic compartment that processes the neurotransmitter signal. During decades of research we have acquired a wealth of knowledge regarding the mechanisms of neurotransmitter release and information processing in the postsynaptic compartment. In great contrast, we know surprisingly little about the pathways that direct the formation, transport, and assembly of the complex molecular machines that make up a functional presynapse.
 We have discovered that presynaptic biogenesis depends on axonal co-transport of synaptic vesicle (SV) and active zone (AZ) proteins in presynaptic lysosome-related vesicles (PLVs). Loss of the lysosomal kinesin adaptor Arl8 results in the accumulation of SV and AZ protein-containing vesicles in neuronal cell bodies and a corresponding depletion of SV and AZ components from presynaptic sites leading to impaired neurotransmission. Conversely, presynaptic function is facilitated upon overexpression of Arl8. We thus postulate an important function for a lysosome-related organelle as an important building block for presynaptic biogenesis (Vukoja et al., Neuron 2018) (Figure 1).

In current work we aim to identify the origin and composition of SV and AZ precursors, dissect the mechanisms of their axonal transport and integration into developing synapses and unravel the pathway that controls axonal transport and presynaptic assembly of newly made SV and AZ proteins to set synaptic weight using stem-cell derived human neurons as a main model. This work is n ot only of fundamental importance for our understanding of synapse formation but may also pave the way These studies are not only of fundamental importance for our understanding of synapse formation but may also pave the way for the future development of therapeutics to cure nerve injury or neurological disorders linked to synapse dysfunction.

Synaptic vesicle  exo- and endocytosis and synaptic vesicle reformation

A major focus of the lab has been the dissection of the mechanisms of presynaptic membrane retrieval and synaptic vesicle reformation (Figure 1). This has resulted in several major discoveries. We found that at physiological temperature SV internalization occurs on several time scales, from less than a second to several seconds and is mediated by dynamin, BAR domain proteins, and the actomyosin cytoskeleton and the activity of actin-nucleating formins such as mDia1 (Soykan et al., Neuron 2017) (Figure 2b). Surprisingly, this pathway of SV membrane endocytosis is independent of clathrin and its major adaptor protein 2 (AP-2), key essential proteins in clathrin-mediated endocytosis in non-neuronal cells. Clathrin/AP-2 instead appear to operate predominantly at internal endosome-like vacuoles to regenerate synaptic vesicles but, is dispensable for compensatory membrane retrieval. Loss of AP-2 in vivo causes the accumulation of endosome-like vacuoles and a depletion of synaptic vesicles, resulting in severely impaired neurotransmission and postnatal lethality (Figure 3).

These experimental results together with theoretical modelling provide a conceptual framework for how synapses capitalize on clathrin-independent membrane retrieval via dynamin/ F-actin and clathrin/ AP-2-mediated SV reformation from endosome-like vacuoles to maintain excitability over a broad range of stimulation frequencies (Kononenko et al., Neuron 2014; Kononenko & Haucke, Neuron 2015) (Figure 4). Consistent with this, we found that mutation of AP-2 causes developmental and epileptic encephalopathy in humans (Helbig et al., Am. J. Hum. Genet. 2019). Our studies highlight the importance of proper AP-2 function for SV recycling (Lopez-Hernandez et al., eLife 2022) and excitatory/ inhibitory balance in the central nervous system.

We have identified specific endocytic adaptors for SV protein sorting such as stonin 2, AP180, and CALM that control the fidelity of SV protein sorting (Kononenko et al., Proc. Natl. Acad. Sci.USA 2013; Koo et al.,Neuron 2015). Moreover, we have unravelled mechanisms that couple vesicle exocytosis and recycling. For example, we have found that loss of the endocytic proteins AP-2 or intersectin 1 in mice causes selective defects in fast neurotransmitter release that may be related to the clearance of release sites for neurotransmitter rather than defects in presynaptic membrane retrieval (Jung, Maritzen et al., EMBO J. 2015, Sakaba et al., Proc. Natl. Acad. Sci. USA 2013., Jaepel et al., Cell Reports 2020). 

Finally, we analyse how axonal and presynaptic components turnover. We have uncovered a surprising function of neuronal autophagy in the the control of the axonal endoplasmic reticulum (ER) and, thereby, in calcium-regulated neurotransmission (Kuijpers et al., Neuron 2021). Present studies aim at addressing the mechanisms and components that regulate axonal ER-phagy in central nervous system neurons. These studies are of importance for the development of novel strategies to combat neurodegeneration and aging-associated memory decline.

Selected publications


  • Lopez- Hernandez, T., Takenaka, K., Mori, Y., Kongpracha, P., Nagamori, S., Haucke, V.*, Takamori, S.* (2022) Clathrin-independent endocytic retrieval of SV proteins mediated by the clathrin adaptor AP-2 at mammalian central synapses. eLife, 11:e71198. [*co-corresponding last authors]
  • Kuijpers, M., Kochlamazashvili, G., Stumpf, A., Puchkov, D., Swaminathan, A., Lucht, M.T., Krause, E., Schmitz, D., Haucke, V. (2021) Neuronal autophagy regulates presynaptic neurotransmission by controlling the axonal endoplasmic reticulum. Neuron 109, 299-313
  • Rizalar, F. S., Roosen, D., Haucke, V. (2021) A presynaptic perspective on transport and assembly mechanisms for synapse formation. N euron 109, 27-41
  • Vukoja A., Rey, U., Petzoldt, A.G., Ott, C., Vollweiter, D., Quentin, C., Puchkov, D., Reynolds, E., Lehmann, M., Hohensee, S., Rosa, S., Lipowsky, R., Sigrist, S., Haucke, V. (2018) Presynaptic biogenesis requires axonal transport of lysosome-related vesicles. Neuron, 99, 1216-1232.
  • Soykan, T., Kaempf, N., Sakaba, T., Vollweiter, D., Goerdeler, F., Puchkov, D., Kononenko, N.L., Haucke, V. (2017) Synaptic vesicle endocytosis occurs on multiple timescales and is mediated by formin-dependent actin assembly. Neuron, 93, 854-866
  • Kononenko & Haucke (2015) Molecular mechanisms of presynaptic membrane retrieval and synaptic vesicle reformation. Neuron, 85, 484-496.
  • Kononenko, N.L., Puchkov, D., Classen, G.A., Walter, A., Pechstein, A., Sawade, L., Kaempf, N., Trimbuch, T., Lorenz, D., Rosenmund, C., Maritzen, M., Haucke, V. (2014) Clathrin/ AP-2 mediate synaptic vesicle reformation from endosome-like vacuoles but are not essential for membrane retrieval at central synapses. Neuron, 82, 981-988.
  • Haucke, V., Neher, E., Sigrist, S.J. (2011) Protein scaffolds in the coupling of synaptic exocytosis and endocytosis. Nat Rev. Mol. Neurosci., 12, 127-138.