Glutamatergic transmission at mossy fiber (MF) synapses on CA3 pyramidal neurons in the hippocampus is mediated by AMPA, kainate, and NMDA receptors and undergoes presynaptic modulation by metabotropic glutamate receptors. The recruitment of different receptors has thus far been studied by altering presynaptic stimulation to modulate glutamate release and interfering pharmacologically with receptors and transporters. Here, we introduce two novel experimental manipulations that alter the fate of glutamate molecules following release. First, an enzymatic glutamate scavenger reduces the postsynaptic response as well as presynaptic modulation by metabotropic receptors. At physiological temperature, however, the scavenger is effective only when glutamate uptake is blocked, revealing a role of active transport in both synaptic and extrasynaptic communication. Second, AMPA and kainate receptor-mediated postsynaptic signals are enhanced when extracellular diffusion is retarded by adding dextran to the perfusion solution, as is feedback modulation by metabotropic receptors, suggesting that the receptors are not saturated under baseline conditions. These results show that manipulating the spatiotemporal profile of glutamate following exocytosis can alter the involvement of different receptors in synaptic transmission.

Acid-sensing ion channel 3 (ASIC3) is involved in acid nociception, but its possible role in neurosensory mechanotransduction is disputed. We report here the generation of Asic3-knockout/eGFPf-knockin mice and subsequent characterization of heterogeneous expression of ASIC3 in the dorsal root ganglion (DRG). ASIC3 is expressed in parvalbumin (Pv+) proprioceptor axons innervating muscle spindles. We further generate a floxed allele of Asic3 (Asic3f/f) and probe the role of ASIC3 in mechanotransduction in neurite-bearing Pv+ DRG neurons through localized elastic matrix movements and electrophysiology. Targeted knockout of Asic3 disrupts spindle afferent sensitivity to dynamic stimuli and impairs mechanotransduction in Pv+ DRG neurons because of substrate deformation-induced neurite stretching, but not to direct neurite indentation. In behavioural tasks, global knockout (Asic3−/−) and Pv-Cre::Asic3f/f mice produce similar deficits in grid and balance beam walking tasks. We conclude that, at least in mouse, ASIC3 is a molecular determinant contributing to dynamic mechanosensitivity in proprioceptors.

The objective of this study was to evaluate the role of ␤-adrenergic receptors in modulating associative long-term potentiation (LTP) induced at CA1 synapses. Two independent Schaffer collateral pathways were stimulated in hippocampal slices. The field EPSP (fEPSP) response evoked in one pathway (the weak pathway) was small, whereas a large response, usually 80 -90% of the maximum, was evoked in the strong pathway. After recording of the baseline fEPSP evoked at 0.033 Hz, LTP of the weak pathway could be associatively induced by paired stimulation of the weak and strong pathways 100 times at 6 sec intervals, with stimulation of the weak pathway preceded 3-10 msec. However, pairing protocols with an interval between stimulation of the two pathways Ͼ10 msec resulted in no LTP. The induced LTP was NMDA receptor dependent, because 50 M D,L-APV blocked its induction. Bath application of 1 M isoproterenol enhanced LTP by increasing the window of the stimulation interval up to 15 msec but did not affect the magnitude of the LTP induced by pairing protocols with intervals Ͻ10 msec. Similar results were obtained when the experiments were repeated using whole-cell recording. These results suggest that activation of ␤-adrenergic receptors can enhance associative LTP by increasing the width of the time window rather than the magnitude of the LTP. Enhancement of LTP by ␤-adrenergic receptors was blocked in slices by pretreatment with inhibitors of protein kinase A or mitogen-activated protein kinase, suggesting that these signaling cascades are involved in this process.

Neuronal transplantation has provided a promising approach for treating neurodegenerative diseases. Recently, efforts have been directed at in vitro induction of various stem cells to transform into neurons. We report the first successful quantities in an in vitro attempt at directing the transformation into neurons of human umbilical mesenchymal cells, which are capable of rapid proliferation in vitro and are easily available. When cultured in neuronal conditioned medium, human umbilical mesenchymal cells started to express neuron-specific proteins such as NeuN and neurofilament (NF) on the 3rd day and exhibited retraction of the cell body, elaboration of processes, clustering of cells and expression of functional mRNA responsible for the synthesis of subunits of the kainate receptor and glutamate decarboxylase on the 6th day. Between the 9th and 12th days, the percentage of human umbilical mesenchymal cells expressing NF was as high as 87%, while functionality was demonstrated by glutamate invoking an inward current. At this stage, cells were differentiated into mature neurons in the post mitosis phase.

Long-term potentiation (LTP) of excitatory transmission is an important candidate cellular mechanism for the storage of memories in the mammalian brain. The subcellular phenomena that underlie the persistent increase in synaptic strength, however, are incompletely understood. A potentially powerful method to detect a presynaptic increase in glutamate release is to examine the effect of LTP induction on the rate at which the use-dependent blocker MK-801 attenuates successive N-methyl-D-aspartic acid (NMDA) receptor-mediated synaptic signals. This method, however, has given apparently contradictory results when applied in hippocampal CA1. The inconsistency could be explained if NMDA receptors were opened by glutamate not only released from local presynaptic terminals, but also diffusing from synapses on neighboring cells where LTP was not induced. Here we examine the effect of pairing-induced LTP on the MK-801 blocking rate in two afferent inputs to dentate granule cells. LTP in the medial perforant path is associated with a significant increase in the MK-801 blocking rate, implying a presynaptic increase in glutamate release probability. An enhanced MK-801 blocking rate is not seen, however, in the lateral perforant path. This result still could be compatible with a presynaptic contribution to LTP in the lateral perforant path if intersynaptic cross-talk occurred. In support of this hypothesis, we show that NMDA receptors consistently sense more quanta of glutamate than do ␣-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors. In the medial perforant path, in contrast, there is no significant difference in the number of quanta mediated by the two receptors. These results support a presynaptic contribution to LTP and imply that differences in intersynaptic cross-talk can complicate the interpretation of experiments designed to detect changes in transmitter release.Long-term potentiation (LTP) is the foremost candidate cellular substrate for memory formation and also may play a major role in the development of neuronal circuits and in neuronal injury (1). In spite of intense effort, the underlying cellular mechanisms of expression remain a matter of some debate (2-4). A potentially sensitive method to detect an increase in presynaptic glutamate release probability is to examine the effect of LTP induction on the rate at which the use-dependent blocker MK-801 attenuates successive excitatory postsynaptic currents (EPSCs) mediated by N-methyl-D-aspartic acid (NMDA) receptors (5, 6). If glutamate is released on a higher proportion of trials after LTP induction, the NMDA receptor-gated channels should be activated more frequently, and MK-801 therefore should produce a faster reduction in the size of the synaptic signal. This method, however, has given conflicting results when applied in the CA1 region of the hippocampus. LTP induced by pairing low-frequency presynaptic stimulation with postsynaptic depolarization was associated with no significant increase in blocking rate (7). Significantly enhanced a...