The dogma that the synaptic cleft acidifies during neurotransmission is based on the co-release of neurotransmitters and protons from synaptic vesicles, and is supported by direct data from sensory ribbon-type synapses. However, it is unclear whether acidification occurs at non-ribbon type synapses. Here we used genetically-encoded fluorescent pH indicators to examine cleft pH at conventional neuronal synapses. At the neuromuscular junction of female Drosophila larvae, we observed alkaline spikes of over 1 log unit during fictive locomotion in vivo Ex vivo, single action potentials evoked alkalinizing pH transients of only ∼0.01 log unit, but these transients summated rapidly during burst firing. A chemical p... More
The dogma that the synaptic cleft acidifies during neurotransmission is based on the co-release of neurotransmitters and protons from synaptic vesicles, and is supported by direct data from sensory ribbon-type synapses. However, it is unclear whether acidification occurs at non-ribbon type synapses. Here we used genetically-encoded fluorescent pH indicators to examine cleft pH at conventional neuronal synapses. At the neuromuscular junction of female Drosophila larvae, we observed alkaline spikes of over 1 log unit during fictive locomotion in vivo Ex vivo, single action potentials evoked alkalinizing pH transients of only ∼0.01 log unit, but these transients summated rapidly during burst firing. A chemical pH indicator targeted to the cleft corroborated these findings. Cleft pH transients were dependent on Ca2+ movement across the postsynaptic membrane, rather than neurotransmitter release per se, a result consistent with cleft alkalinization being driven by the Ca2+/H+ antiporting activity of the plasma membrane Ca2+-ATPase at the postsynaptic membrane. Targeting the pH indicators to the microenvironment of the presynaptic voltage-gated Ca2+ channels revealed that alkalinization also occurred within the cleft proper at the active zone and not just within extra-synaptic regions. Application of the pH indicators at the mouse calyx of Held, a mammalian central synapse, similarly revealed cleft alkalinization during burst firing in both males and females. These findings, made at two quite different non-ribbon type synapses, suggest that cleft alkalinization during neurotransmission, rather than acidification, is a generalizable phenomenon across conventional neuronal synapses.SIGNIFICANCE STATEMENTNeurotransmission is highly sensitive to the pH of the extracellular milieu. This is readily evident in the neurological symptoms that accompany systemic acid/base imbalances. Imaging data from sensory ribbon-type synapses show that neurotransmission itself can acidify the synaptic cleft, likely due to the co-release of protons and glutamate. It is not clear if the same phenomenon occurs at conventional neuronal synapses due to the difficulties in collecting such data. If it does occur, it would provide for an additional layer of activity-dependent modulation of neurotransmission. Our findings of alkalinization, rather than acidification, within the cleft of two different neuronal synapses encourages a reassessment of the scope of activity-dependent pH influences on neurotransmission and short-term synaptic plasticity.,Copyright © 2020 the authors.
