Jane M. Sullivan

Synaptic vesicle protein 2 enhances release probability at quiescent synapses

We report a thorough analysis of neurotransmission in cultured hippocampal neurons lacking synaptic vesicle protein 2 (SV2), a membrane glycoprotein present in all vesicles that undergo regulated secretion. We found that SV2 selectively enhances low-frequency neurotransmission by priming morphologically docked vesicles. Loss of SV2 reduced initial release probability during a train of action potentials but had no effect on steady-state responses. The amount and decay rate of asynchronous release, two measures sensitive to presynaptic calcium concentrations, are not altered in SV2 knock-outs, suggesting that SV2 does not act by modulating presynaptic calcium. Normal neurotransmission could be temporarily recovered by delivering an exhaustive stimulus train. Our results indicate that SV2 primes vesicles in quiescent neurons and that SV2 function can be bypassed by an activity-dependent priming mechanism. We propose that SV2 action modulates synaptic networks by ensuring that low-frequency neurotransmission is faithfully conveyed.

Amyloid precursor protein overexpression depresses excitatory transmission through both presynaptic and postsynaptic mechanisms

Overexpression of the amyloid precursor protein (APP) in hippocampal neurons leads to elevated β-amyloid peptide (Aβ) production and consequent depression of excitatory transmission. The precise mechanisms underlying APP-induced synaptic depression are poorly understood. Uncovering these mechanisms could provide insight into how neuronal function is compromised before cell death during the early stages of Alzheimers disease. Here we verify that APP up-regulation leads to depression of transmission in cultured hippocampal autapses; and we perform whole-cell recording, FM imaging, and immunocytochemistry to identify the specific mechanisms accounting for this depression. We find that APP overexpression leads to postsynaptic silencing through a selective reduction of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor-mediated currents. This effect is likely mediated by Aβ because expression of mutant APP incapable of producing Aβ did not depress transmission. In addition, although we eliminate presynaptic silencing as a mechanism underlying APP-mediated inhibition of transmission, we did observe an Aβ-induced presynaptic deficit in vesicle recycling with sustained stimulation. These findings demonstrate that APP elevation disrupts both presynaptic and postsynaptic compartments.

ASCL1 reprograms mouse Müller glia into neurogenic retinal progenitors

Non-mammalian vertebrates have a robust ability to regenerate injured retinal neurons from Müller glia (MG) that activate the gene encoding the proneural factor Achaete-scute homolog 1 (Ascl1; also known as Mash1 in mammals) and de-differentiate into progenitor cells. By contrast, mammalian MG have a limited regenerative response and fail to upregulate Ascl1 after injury. To test whether ASCL1 could restore neurogenic potential to mammalian MG, we overexpressed ASCL1 in dissociated mouse MG cultures and intact retinal explants. ASCL1-infected MG upregulated retinal progenitor-specific genes and downregulated glial genes. Furthermore, ASCL1 remodeled the chromatin at its targets from a repressive to an active configuration. MG-derived progenitors differentiated into cells that exhibited neuronal morphologies, expressed retinal subtype-specific neuronal markers and displayed neuron-like physiological responses. These results indicate that a single transcription factor, ASCL1, can induce a neurogenic state in mature MG.

Amyloid-β1–42 Slows Clearance of Synaptically Released Glutamate by Mislocalizing Astrocytic GLT-1

GLT-1, the major glutamate transporter in the adult brain, is abundantly expressed in astrocytic processes enveloping synapses. By limiting glutamate escape into the surrounding neuropil, GLT-1 preserves the spatial specificity of synaptic signaling. Here we show that the amyloid-β peptide Aβ1–42 markedly prolongs the extracellular lifetime of synaptically released glutamate by reducing GLT-1 surface expression in mouse astrocytes and that this effect is prevented by the vitamin E derivative Trolox. These findings indicate that astrocytic glutamate transporter dysfunction may play an important role in the pathogenesis of Alzheimers disease and suggest possible mechanisms by which several current treatment strategies could protect against the disease.

MicroRNA132 Modulates Short-Term Synaptic Plasticity but Not Basal Release Probability in Hippocampal Neurons

MicroRNAs play important regulatory roles in a broad range of cellular processes including neuronal morphology and long-term synaptic plasticity. MicroRNA-132 (miR132) is a CREB-regulated miRNA that is induced by neuronal activity and neurotrophins, and plays a role in regulating neuronal morphology and cellular excitability. Little is known about the effects of miR132 expression on synaptic function. Here we show that overexpression of miR132 increases the paired-pulse ratio and decreases synaptic depression in cultured mouse hippocampal neurons without affecting the initial probability of neurotransmitter release, the calcium sensitivity of release, the amplitude of excitatory postsynaptic currents or the size of the readily releasable pool of synaptic vesicles. These findings are the first to demonstrate that microRNAs can regulate short-term plasticity in neurons.

Presenilin 1 regulates homeostatic synaptic scaling through Akt signaling

Neurons adapt to long-lasting changes in network activity, both in vivo and in vitro, by adjusting their synaptic strengths to stabilize firing rates. We found that homeostatic scaling of excitatory synapses was impaired in hippocampal neurons derived from mice lacking presenilin 1 (Psen1−/− mice) or expressing a familial Alzheimers disease–linked Psen1 mutation (Psen1M146V). These findings suggest that deficits in synaptic homeostasis may contribute to brain dysfunction in Alzheimers disease.

Synaptotagmin IV Does Not Alter Excitatory Fast Synaptic Transmission or Fusion Pore Kinetics in Mammalian CNS Neurons

Synaptotagmin IV (Syt IV) is a brain-specific isoform of the synaptotagmin family, the levels of which are strongly elevated after seizure activity. The dominant hypothesis of Syt IV function states that Syt IV upregulation is a neuroprotective mechanism for reducing neurotransmitter release. To test this hypothesis in mammalian CNS synapses, Syt IV was overexpressed in cultured mouse hippocampal neurons, and acute effects on fast excitatory neurotransmission were assessed. We found neurotransmission unaltered with respect to basal release probability, Ca2+ dependence of release, short-term plasticity, and fusion pore kinetics. In contrast, expression of a mutant Syt I with diminished Ca2+ affinity (R233Q) reduced release probability and altered the Ca2+ dependence of release, thus demonstrating the sensitivity of the system to changes in neurotransmission resulting from changes to the Ca2+ sensor. Together, these data refute the dominant model that Syt IV functions as an inhibitor of neurotransmitter release in mammalian neurons.

NO going back

After exocytosis, synaptic vesicles must be retrieved and refilled with neurotransmitter to supply the needs of an active neuron. A new report finds that synaptic activity, through the retrograde action of nitric oxide (NO), regulates the rate of this synaptic vesicle recycling. These findings suggest that NOmight enhance the synaptic strength of coincidentally active neurons.

Synaptotagmin mutants Y311N and K326/327A alter the calcium dependence of neurotransmission

Synaptotagmin I, a calcium-binding synaptic vesicle protein, is thought to act as the calcium sensor for fast neurotransmission, but what synaptotagmin I does, upon binding calcium, to trigger exocytosis is still unknown. To begin to examine the role of synaptotagmin Is interactions with calcium-dependent binding partners, three mutant versions of synaptotagmin I reported to affect calcium-dependent self-oligomerization (Y311N, K327A, and K326/327A) were expressed in cultured mouse hippocampal neurons lacking endogenous synaptotagmin I, and effects on neurotransmission were evaluated by comparison with transmission rescued by wild-type synaptotagmin I. All three mutants reduced transmitter release. To separate effects on calcium binding from effects on calcium-dependent oligomerization, we measured the calcium dependence of exocytosis for two of the mutants. Both showed apparent calcium affinity much lower than wild-type, a reduction sufficient to account for the neurotransmission defects. We conclude that self-oligomerization is unlikely to play any significant role in triggering synaptic vesicle exocytosis.

Calcium sensor regulation of the CaV2.1 Ca2+ channel contributes to short-term synaptic plasticity in hippocampal neurons

Significance Information processing in the brain is mediated through synaptic connections between neurons, where neurotransmitter molecules released from presynaptic nerve terminals stimulate postsynaptic cells. Strength of synaptic transmission is increased transiently by short-term synaptic facilitation in response to repeated stimulation of nerve fibers. Synaptic transmission is initiated by calcium influx through calcium channels in presynaptic nerve terminals, which are regulated by calcium sensor proteins. We found that genetically modified mice in which we introduced a mutation in the binding site for calcium sensor proteins in presynaptic calcium channels have substantially altered facilitation in hippocampal synapses in response to pairs or trains of repetitive high-frequency stimuli. Our results show that disruption of calcium channel regulation by calcium sensor proteins impairs short-term facilitation in native synapses. Short-term synaptic plasticity is induced by calcium (Ca2+) accumulating in presynaptic nerve terminals during repetitive action potentials. Regulation of voltage-gated CaV2.1 Ca2+ channels by Ca2+ sensor proteins induces facilitation of Ca2+ currents and synaptic facilitation in cultured neurons expressing exogenous CaV2.1 channels. However, it is unknown whether this mechanism contributes to facilitation in native synapses. We introduced the IM-AA mutation into the IQ-like motif (IM) of the Ca2+ sensor binding site. This mutation does not alter voltage dependence or kinetics of CaV2.1 currents, or frequency or amplitude of spontaneous miniature excitatory postsynaptic currents (mEPSCs); however, synaptic facilitation is completely blocked in excitatory glutamatergic synapses in hippocampal autaptic cultures. In acutely prepared hippocampal slices, frequency and amplitude of mEPSCs and amplitudes of evoked EPSCs are unaltered. In contrast, short-term synaptic facilitation in response to paired stimuli is reduced by ∼50%. In the presence of EGTA-AM to prevent global increases in free Ca2+, the IM-AA mutation completely blocks short-term synaptic facilitation, indicating that synaptic facilitation by brief, local increases in Ca2+ is dependent upon regulation of CaV2.1 channels by Ca2+ sensor proteins. In response to trains of action potentials, synaptic facilitation is reduced in IM-AA synapses in initial stimuli, consistent with results of paired-pulse experiments; however, synaptic depression is also delayed, resulting in sustained increases in amplitudes of later EPSCs during trains of 10 stimuli at 10–20 Hz. Evidently, regulation of CaV2.1 channels by CaS proteins is required for normal short-term plasticity and normal encoding of information in native hippocampal synapses.