Energy demand of synaptic transmission at the hippocampal Schaffer-collateral synapse

Abstract

Neuroenergetic models of synaptic transmission predicted that energy demand is highest for action potentials (APs) and postsynaptic ion fluxes, whereas the presynaptic contribution is rather small. Here, we addressed the question of energy consumption at Schaffer-collateral synapses. We monitored stimulus-induced changes in extracellular potassium, sodium, and calcium concentration while recording partial oxygen pressure (pO(2)) and NAD(P)H fluorescence. Blockade of postsynaptic receptors reduced ion fluxes as well as pO(2) and NAD(P)H transients by ∼50%. Additional blockade of transmitter release further reduced Na(+), K(+), and pO(2) transients by ∼30% without altering presynaptic APs, indicating considerable contribution of Ca(2+)-removal, transmitter and vesicle turnover to energy consumption.

Figure 1

Changes in field potential (fp) responses and excitatory postsynaptic potential amplitude (EPSPs) during sequential blockade of postsynaptic receptors, voltage-gated Ca²⁺-channels, and action potentials (APs). (A) Schematic representation of the recording design and stimulus-induced fp responses in the str. radiatum (SR) of CA1 (SP, stratum pyramidale and SL: stratum lacunosum moleculare). Presynaptic APs were represented by the afferent volley, the shortest latency component of the fp response. Postsynaptic responses including postsynaptic population spike and field EPSPs (fEPSP) were quantified by measuring the amplitude of the subsequent negative deflection of the fp response. (B) Effect of sequential blockade of glutamate and GABA_A receptors and APs on the different components of the fp response in percentage of the control (CTL). Note that the cocktail blocks fEPSP and postsynaptic APs completely within 20 minutes. Field potential is completely abolished in the presence of TTX, indicating that no direct activation of postsynaptic cells occurred. (C) Effect of sequential blockade of glutamate and GABA_A receptors and presynaptic Ca²⁺-entry by Ni²⁺ on the different components of the fp response in percentage of the control. Note that presynaptic APs remain unchanged in the presence of Ni²⁺. (D) Effect of sequential blockade of glutamate and GABA_A receptors and presynaptic Ca²⁺-entry by ω-Conotoxin MVIIC/ω-Agatoxin TK (ω-Con/ω-Aga) on the different components of the fp response in percentage of the control. Similarly to Ni²⁺ ω-Con/ω-Aga did not influence presynaptic APs as represented by the afferent volley. (E) Effect of glutamate and GABA_A receptors on the EPSCs/IPSCs in CA1 pyramidal cell under submerged conditions. Overlay of EPSCs/IPSCs of a stimulus train aligned by the stimulation artifact (upper traces). Note the disappearance of synaptic components in the presence of the cocktail. First six pulses of stimulus train as recorded with a sharp electrode from a CA1 pyramidal cell under interface conditions (lower trace). Note a slight increase of the EPSP amplitude from the first to the second stimulus. (F) Overlay of EPSPs during sequential blockade of glutamate and GABA_A receptors and presynaptic Ca²⁺-entry by Ni²⁺. EPSP was almost completely inhibited by the cocktail, whereas subsequent application of Ni²⁺ did not cause a further reduction of the EPSP amplitude.

Figure 2

Kinetics of stimulus-induced [K⁺]_o, [Ca²⁺]_o, [Na⁺]_o, partial oxygen pressure (pO₂), and NAD(P)H transients. (A) Simultaneously measured changes in [K⁺]_o, [Na⁺]_o, and pO₂. The start of the decrease in pO₂ is delayed as compared with the ion fluxes and the falling phase of the pO₂ transient overlaps with the phase of recovery of ion transients. (B) Simultaneously measured changes in [K⁺]_o, [Ca²⁺]_o, and pO₂. Although recovery of [Ca²⁺]_o is slightly slower than that of [Na⁺]_o, it still overlaps with the largest part of falling phase of pO₂. (C) Simultaneous recording of pO₂ and NAD(P)H fluorescence transients under submerged conditions. Decrease of the NAD(P)H fluorescence represents a shift to the oxidized, an increase in the fluorescence to the reduced form of NAD(P)H. pO₂ as measured with the Clark-style oxygen electrode lags behind the changes in the fluorescence, suggesting that O₂ diffusion in the tissue might alter the kinetics of pO₂ transients. Note that NAD(P)H/NAD(P)⁺ ratio is increased during the whole period of enhanced O₂ consumption. Stimulation lengths are indicated with the bars on the top of (A–C). (D) Amplitude of stimulus-induced pO₂ and [K⁺]_o transients did not change significantly over 100 minutes of repeated stimulation, indicating that neither synaptic facilitation nor synaptic depression occur with 20 Hz stimulation protocol.

Figure 3

Ion fluxes, oxygen consumption, and redox changes as created by pre- and postsynaptic processes. Changes in [K⁺]_o (A), [Ca²⁺]_o (B), [Na⁺]_o (C), and partial oxygen pressure (pO₂) (D) transients during sequential blockade of glutamate and GABA_A receptors and action potentials (APs). (E) Comparison of the changes in ion fluxes and oxygen consumption in percentage of control. Note that ∼50% of ion fluxes and pO₂ changes were removed by inhibition of glutamate and GABAergic transmission. Asterisks mark statistical significance (repeated measures analysis of variance (ANOVA)). (Fa) Changes in biphasic NAD(P)H transients during blockade of glutamatergic and GABAergic transmission, the insert representing the area where the fluorescence was recorded. (Fb) Comparison of dip and overshoot components of NAD(P)H transients. Both components decreased equally in the presence of the cocktail.

Figure 4

Ion fluxes, oxygen consumption, and redox changes as created by presynaptic action potentials (APs) and transmitter release linked processes. Changes in [K⁺]_o (A), [Ca²⁺]_o (B), [Na⁺]_o (C), and partial oxygen pressure (pO₂) (D) transients during sequential blockade of glutamate and GABA_A receptors and vesicle release by Ni²⁺. (E) Comparison of the changes in ion fluxes and oxygen consumption in percentage of control. Note that ∼50% of ion fluxes and pO₂ changes were removed by inhibition of glutamate and GABAergic transmission and Ni²⁺ further decreased all ion and pO₂ transients by ∼30%, in spite of the unaltered presynaptic APs. Asterisks mark statistical significance (repeated measures analysis of variance (ANOVA)). (Fa) Changes in biphasic NAD(P)H transients during blockade of glutamatergic and GABAergic transmission and vesicle release by Ni²⁺. (Fb) Comparison of dip and overshoot components of NAD(P)H transients. Both components were further decreased in the presence of Ni²⁺.

Figure 5

Changes in stimulus-induced [K⁺]_o and partial oxygen pressure (pO₂) transients during sequential blockade of synaptic transmission and presynaptic Ca²⁺-entry. Changes in [K⁺]_o (A) and pO₂ (B) transients during sequential blockade of glutamate and GABA_A receptors and vesicle release by ω-Conotoxin MVIIC/ω-Agatoxin TK (ω-Con/ω-Aga). (C) Comparison of the changes in [K⁺]_o fluxes and oxygen consumption in percentage of control. ω-Con/ω-Aga decreased [K⁺]_o and pO₂ transients by about an additional 30%. Asterisks mark statistical significance (repeated measures analysis of variance (ANOVA)). (D) Comparison of the effects of Ni²⁺ and ω-Con/ω-Aga on [K⁺]_o and pO₂ transients. Although Ni²⁺ had a significantly larger effect on the [K⁺]_o transients, the reduction in pO₂ transients was not different between Ni²⁺ and ω-Con/ω-Aga-treated groups (Student's t-test).