At hippocampal CA1 synapses, a single release event may be associated with the
release of up to 10 vesicles [58]. In particular, an individual mossy fiber axon has
approximately 15 giant boutons (3–5 μm diameter at 150 μm intervals) and 37 release
sites (active zones) each [51, 52, 59]. Single boutons contain about 20 active zones at
0.45 μm intervals and 16,000 vesicles, being 1400 ready for release that may be
multivesicular, thus leading to very intense release processes [51, 60]. It has been
reported that the free or loosely bound zinc and glutamate vesicular concentrations
are 1–5 and 60–210 mM, respectively [16, 27, 47]. Let’s assume that zinc and glutamate, which have similar diffusion coefficients [20, 27], are co-released in the same
proportion and that, as what happens for glutamate, the cleft released zinc concentration becomes uniform in tens of microseconds [16]. It can be estimated that the
volume of a mossy fiber-CA3 cleft (20 nm width, 3 μm radius) is much higher
(approximately 17,000 times) than that of a vesicle (40 nm diameter) [27, 49–51,
61, 62]. The cleft real volume is significantly smaller, by about 25%, if the volume of
densely packed conic (20 nm height) dendritic protrusions is subtracted [63]. In this
case the discharge of an individual vesicle would lead to the following initial concentrations: 80–400 nM for zinc and 4–20 μM for glutamate. There is a large difference
between the radii and, thus, the volumes of the mossy fiber-CA3 and the CA3-CA1
clefts since the latter has a 20 nm width and only 250 nm radius. As a consequence, in
the CA1 region, the cleft glutamate range of concentrations is estimated to be around
0.4–2 mM and is thus close to the previously reported ranges of values, 1–5 mM [16]
and 0.25–11 mM following an individual vesicle release [64]. Another important issue
is the role of the connection between the cleft and extrasynaptic regions with much
higher volume [65]. This volume has to be added to that of the cleft, since it forms a
large part of the space where the zinc concentration changes occur. A similar fact has
been considered for cleft glutamate clearance in neurons of the central nervous
system, where the glutamate concentration decreased very rapidly (1–5 ms) 100–500
times [65]. Let us assume again that zinc and glutamate diffuse in a similar way
[20, 27]. In this case the concentration range of cleft free zinc changes, evoked by an
individual vesicle discharge and after diffusing away from the cleft, will be 0.8–4 nM.
If multivesicular release occurs [60], the amount of zinc in the cleft will be significantly larger.
Another estimate of cleft zinc discharge and uptake can be obtained from fluorescent glutamate signals associated with single or repetitive stimulation applied to
cultured hippocampal neurons [66]. These authors have found that for the single
and short types of stimulation, the maximum concentrations of glutamate were
around 0.3 and 0.8 μM, respectively, occurring clearance in less than 1 s, for the
single, and 2 s, for the short stimuli. If, as previously estimated, there is about 50
times more glutamate in the cleft, the equivalent maximum concentration range for
free zinc will be 6–16 nM.
Previous work has reported that the resting free zinc concentration in the cleft is
below 10 nM, meaning that the NMDA 2A sites, which are highly sensitive for zinc,
will not become saturated by zinc, and also that the amount of zinc discharged by
an individual stimulus does not seem to alter much postsynaptic NMDA currents
[27]. Our estimates, for more intense stimulations such as the short and long processes, which are associated with 10 and 100 times more free zinc in the cleft,
respectively, than for the single process, indicate that the NMDA 2A sites
(K D = 6 nM) are nearly or fully occupied as reported earlier [10, 27, 28]. For the
short and long stimulations, the amount of GLAST complexes is approximately 2
and 10 times more than for NMDA 2A complexes, respectively. Also, only for these
stronger stimulation protocols, the much lower-affinity NMDA 2B and AMPA
receptors become significantly occupied. The K ATP channels and the EAAT4 transporters form a reasonable amount of zinc complexes in spite of their smaller
123
Computer Simulations of Hippocampal Mossy Fiber Cleft Zinc Movements
DOI: http://dx.doi.org/10.5772/intechopen.90094
release of up to 10 vesicles [58]. In particular, an individual mossy fiber axon has
approximately 15 giant boutons (3–5 μm diameter at 150 μm intervals) and 37 release
sites (active zones) each [51, 52, 59]. Single boutons contain about 20 active zones at
0.45 μm intervals and 16,000 vesicles, being 1400 ready for release that may be
multivesicular, thus leading to very intense release processes [51, 60]. It has been
reported that the free or loosely bound zinc and glutamate vesicular concentrations
are 1–5 and 60–210 mM, respectively [16, 27, 47]. Let’s assume that zinc and glutamate, which have similar diffusion coefficients [20, 27], are co-released in the same
proportion and that, as what happens for glutamate, the cleft released zinc concentration becomes uniform in tens of microseconds [16]. It can be estimated that the
volume of a mossy fiber-CA3 cleft (20 nm width, 3 μm radius) is much higher
(approximately 17,000 times) than that of a vesicle (40 nm diameter) [27, 49–51,
61, 62]. The cleft real volume is significantly smaller, by about 25%, if the volume of
densely packed conic (20 nm height) dendritic protrusions is subtracted [63]. In this
case the discharge of an individual vesicle would lead to the following initial concentrations: 80–400 nM for zinc and 4–20 μM for glutamate. There is a large difference
between the radii and, thus, the volumes of the mossy fiber-CA3 and the CA3-CA1
clefts since the latter has a 20 nm width and only 250 nm radius. As a consequence, in
the CA1 region, the cleft glutamate range of concentrations is estimated to be around
0.4–2 mM and is thus close to the previously reported ranges of values, 1–5 mM [16]
and 0.25–11 mM following an individual vesicle release [64]. Another important issue
is the role of the connection between the cleft and extrasynaptic regions with much
higher volume [65]. This volume has to be added to that of the cleft, since it forms a
large part of the space where the zinc concentration changes occur. A similar fact has
been considered for cleft glutamate clearance in neurons of the central nervous
system, where the glutamate concentration decreased very rapidly (1–5 ms) 100–500
times [65]. Let us assume again that zinc and glutamate diffuse in a similar way
[20, 27]. In this case the concentration range of cleft free zinc changes, evoked by an
individual vesicle discharge and after diffusing away from the cleft, will be 0.8–4 nM.
If multivesicular release occurs [60], the amount of zinc in the cleft will be significantly larger.
Another estimate of cleft zinc discharge and uptake can be obtained from fluorescent glutamate signals associated with single or repetitive stimulation applied to
cultured hippocampal neurons [66]. These authors have found that for the single
and short types of stimulation, the maximum concentrations of glutamate were
around 0.3 and 0.8 μM, respectively, occurring clearance in less than 1 s, for the
single, and 2 s, for the short stimuli. If, as previously estimated, there is about 50
times more glutamate in the cleft, the equivalent maximum concentration range for
free zinc will be 6–16 nM.
Previous work has reported that the resting free zinc concentration in the cleft is
below 10 nM, meaning that the NMDA 2A sites, which are highly sensitive for zinc,
will not become saturated by zinc, and also that the amount of zinc discharged by
an individual stimulus does not seem to alter much postsynaptic NMDA currents
[27]. Our estimates, for more intense stimulations such as the short and long processes, which are associated with 10 and 100 times more free zinc in the cleft,
respectively, than for the single process, indicate that the NMDA 2A sites
(K D = 6 nM) are nearly or fully occupied as reported earlier [10, 27, 28]. For the
short and long stimulations, the amount of GLAST complexes is approximately 2
and 10 times more than for NMDA 2A complexes, respectively. Also, only for these
stronger stimulation protocols, the much lower-affinity NMDA 2B and AMPA
receptors become significantly occupied. The K ATP channels and the EAAT4 transporters form a reasonable amount of zinc complexes in spite of their smaller
123
Computer Simulations of Hippocampal Mossy Fiber Cleft Zinc Movements
DOI: http://dx.doi.org/10.5772/intechopen.90094