The Role of NMDA Receptor Co-Agonists in Dendritic Integration

Abstract

The integration of synaptic inputs is the fundamental function of neurons. This integration does not only take place at the soma, but also dendrites can actively shape the input-output computations of a neuronal network. In the CA1 region of the hippocampus, dendrites of pyramidal cells integrate inputs either linear or, under specific circumstances, supralinear. For instance, synchronous, spatially clustered inputs at apical oblique dendrites have been shown to evoke dendritic spikes. Likewise, specifically timed activation of Schaffer collateral and perforant path fibers results in large dendritic plateau potentials. These two types of nonlinear integration are highly dependent on NMDA receptor recruitment. Glutamate uptake and dynamic supply of co-agonists can modulate NMDA receptor recruitment. Astrocytes take up the majority of the released glutamate. Moreover, they are one possible source of NMDA receptor co-agonists. In astrocytes, the release of gliotransmitters, such as the NMDA receptor co-agonist D-serine, is evoked by calcium transients. <br /> With my thesis, I aim to unravel the role of astrocytes in different types of dendritic integration through modulation of glutamate uptake or NMDA receptor co-agonist supply. Furthermore, the recruitment and relevance of astrocytic, calcium-dependent, dynamic supply of D-serine under behaviorally relevant activity patterns were explored. Conclusively, the consequence of disrupting dynamic co-agonist supply on learning and memory in vivo was investigated. To this end, whole-cell patch clamp recordings in combination with microiontophoretic glutamate application, electrical stimulation and two-photon excitation fluorescence microscopy as well as behavioral experiments were used. <br /> My data showed that glutamate uptake in the CA1 region of the hippocampus was more tightly regulated at small compared to large spines. Accordingly, the probability of evoking a dendritic spike was increased at the latter. Moreover, my results indicated that local dendritic integration can be modulated by exogenous co-agonists but also through cannabinoid-evoked astrocytic calcium transients that induce the release of endogenous NMDA receptor co-agonist. Endocannabinoids can be mobilized through neuronal depolarization. In my experiments, I therefore used axonal stimulation of CA1 pyramidal cells at different frequencies to investigate whether endogenously mobilized cannabinoids could modulate dendritic integration. Endocannabinoids evoked a similar effect on the integration as exogenous cannabinoids or direct NMDA receptor co-agonist application. I demonstrated that this modulation depends on CB1 receptors and the release of the NMDA receptor co-agonist D-serine. The data showed an unexpected dependency of the frequency of neuronal activation on the dendritic integration. Typically, the mobilization of endocannabinoids is increased with increasing depolarization and action potential number. Interestingly, the opposite was observed. While action potentials evoked at a frequency of 10 Hz increased the threshold and amplitude of dendritic spikes, axonal stimulation at 40 Hz did not. Additional experiments revealed that hyperpolarization-activated cyclic nucleotide-gated (HCN) channels, which are known for the relevance in establishing the neuronal resonance, are required for the frequency-dependent modulation of dendritic integration. Importantly, specific knock-out of CB1 receptors in astrocytes disrupted the positive feedback loop and resulted in selective impairment of spatial reversal learning in a passive place avoidance task and altered long-term object recognition and location memory. Astrocytic calcium transients are the key step in linking neuronal activity to the release of D-serine. To gain a better understanding of these astrocytic calcium transients, the relationship between the pre-event baseline and the peak or amplitude calcium concentration was investigated. Spontaneous and evoked astrocytic calcium transients were monitored in situ and in vivo by two-photon excitation microscopy. These experiments revealed an overall positive relationship between the pre-event baseline and the peak calcium concentration. Contrastingly, the transient’s amplitude and the pre-event baseline concentration correlated negatively with each other. While influx of extracellular calcium mediated the former, the latter was depending on store-dependent calcium release. This relationship poses interesting possibilities on how the intracellular calcium concentration of astrocytes could shape the release of gliotransmitters and synaptic transmission. <br /> My thesis revealed a previously unknown, behaviorally relevant, frequency-dependent positive feedback loop that links pyramidal cell activity to their dendritic integration. Through endocannabinoid mobilization and astrocytic calcium transients, a subsequent increase in D- serine is evoked. D-serine in turn increases the recruitment of NMDA receptors and hence, promotes supralinear dendritic integration through a decreased threshold and increased amplitude of dendritic spikes. The findings not only expand our understanding of astrocytic glutamate clearance and astrocytic calcium transients but also underline the importance of astrocytes for learning and memory through gliotransmission.