Flotillin‐1 promotes formation of glutamatergic synapses in hippocampal neurons

Abstract

Synapse malformation underlies numerous neurodevelopmental illnesses, including autism spectrum disorders. Here we identify the lipid raft protein flotillin-1 as a promoter of glutamatergic synapse formation. We cultured neurons from the hippocampus, a brain region important for learning and memory, and examined them at two weeks in vitro, a time period rich with synapse formation. Double-label immunocytochemistry of native flot-1 with glutamatergic and GABAergic synapse markers showed that flot-1 was preferentially colocalized with the glutamatergic presynaptic marker vesicular glutamate transporter 1 (VGLUT1), compared to the GABAergic presynaptic marker glutamic acid decarboxylase-65 (GAD-65). Triple-label immunocytochemistry of native flot-1, VGLUT1, and NR1, the obligatory subunit of NMDA receptors, indicates that Flot-1 was preferentially localized to synaptic rather than extrasynaptic NR1. Furthermore, electrophysiological results using whole-cell patch clamp showed that Flot-1 increased the frequency of miniature excitatory postsynaptic currents (mEPSCs) but not miniature inhibitory postsynaptic currents (mIPSCs), whereas amplitude and decay kinetics of either type of synaptic current was not affected. Corresponding immunocytochemical data confirmed that the number of glutamatergic synapses increased with flot-1 overexpression. Overall, our anatomical and physiological results show that flot-1 enhances the formation of glutamatergic synapses but not GABAergic synapses, suggesting that the role of flot-1 in neurodevelopmental disorders should be explored.

Reggie-1 and reggie-2 (flotillins) participate in Rab11a-dependent cargo trafficking, spine synapse formation and LTP-related AMPA receptor (GluA1) surface exposure in mouse hippocampal neurons

“…These studies are consistent with the present results and emphasize the role of reggie in brain function and synaptic plasticity. The findings by Swanwick et al (2010) and our own results suggest that reggie-2 may have an even stronger influence on spine formation than reggie-1. Reggie-2 downregulation reduced spine number by 59%, and reggie-1 siRNA by 24%.…”

“…It was reported that reggie-2 overexpression in maturated hippocampal neurons leads to increased spines and EPSPs (Swanwick et al, 2010). Another study showed by mass spectrometry that reggies were also upregulated during memory related strengthening of synapses in Aplysia (Monje et al, 2013) and during barrel cortex plasticity (Butko et al, 2013).…”

“…Flotillin-1/reggie-2 is downregulated in the cortical barrel fields after sensory deprivation (Butko et al, 2013). Upregulation of flotillin-1/reggie-2 has, in addition, been implicated in the formation of glutamatergic synapses of mouse hippocampal neurons in vitro (Swanwick et al, 2010) and increase of the frequency of miniature excitatory postsynaptic currents.…”

The roles of flotillin microdomains – endocytosis and beyond

“…As mentioned above, flotillins also cluster with antibody-patched cell adhesion molecules like Thy1 and F3 in neurons (Lang et al, 1998). In hippocampal neurons, flotillin 1 promotes formation of glutamatergic synapses (Swanwick et al, 2010b) and interacts with synaptic adhesion-like molecules (SALMs) to regulate neurite outgrowth by coordinating adhesion, exocytosis and actin cytoskeleton dynamics (Swanwick et al, 2010a).…”

Molecular and structural bases for postsynaptic signal processing: interaction between postsynaptic density and postsynaptic membrane rafts

“…Flotillin-1, a resident protein in noncaveolar rafts, 25 is localized to excitatory synapses, and its overexpression enhances formation of glutamatergic synapses in hippocampal neurons. 74 Flotillin-1 induces filopodia formation 74 and promotes hippocampal neuronal differentiation. 75 Flotillin induces growth cone formation and neurite outgrowth via an interaction with prion protein.…”