1. Chapter 10 The ionotropic glutamate receptors
From Cellular and Molecular Neurophysiology, Fourth Edition.
Copyright © 2015 Elsevier Ltd. All rights reserved.

2. Figure 10.1 Pharmacology of ionotropic glutamate receptors.
(a) AMPA: α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid; ATPA : (RS)-2-amino-3-(3-hydroxy-5-tert-butylisoxazol-4-yl)propanoic acid selective agonist of GluK1-containing receptors; CNQX : 6-cyano- 7-nitroquinoxaline-2,3-dione; D-AP5 :DL-2-amino-5-phosphonopentanoic acid; DNQX: 6,7-dinitroquinoxaline-2,3-dione; GYKI 52466: 4-(8-methyl-9H-1,3-dioxolo[4,5-h,2,3]benzodiazepin-5-yl)-benzenamine dihydrochloride; GYKI 53655: 1-(4-aminophenyl)-3-methylcarbamyl-4-methyl-3,4-dihydro-7,8-methylenedioxy-5H-2,3-benzodiazepine hydrochloride; MK 801:(5S,10R)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate or dizocilpine hydrogen maleate; NBQX: 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo[f]quinoxaline-7-sulfonamide; SYM 2081: (2S,4R)-4-methylglutamic acid; UBP 310: (S)-1-(2-amino-2-carboxyethyl)-3-(2-carboxy-thiophene-3-yl-methyl)-5-methylpyrimidine-2,4-dione selective antagonist of GluK1/GluK3. (b) Agonists of iGluRs. (c) Family of ionotropic glutamate receptor subunits.
Part (c) adapted from Wollmuth LP, Sobolevsky AI (2004) Structure and gating of the glutamate receptor ion channel. Trend. Neurosci.27, 321–328, with permission.
From Cellular and Molecular Neurophysiology, Fourth Edition.
Copyright © 2015 Elsevier Ltd. All rights reserved.

3. 3 Figure 10.1 Pharmacology of ionotropic glutamate receptors.(cont.)
(a) AMPA: α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid; ATPA : (RS)-2-amino-3-(3-hydroxy-5-tert-butylisoxazol-4-yl)propanoic acid selective agonist of GluK1-containing receptors; CNQX : 6-cyano- 7-nitroquinoxaline-2,3-dione; D-AP5 :DL-2-amino-5-phosphonopentanoic acid; DNQX: 6,7-dinitroquinoxaline-2,3-dione; GYKI 52466: 4-(8-methyl-9H-1,3-dioxolo[4,5-h,2,3]benzodiazepin-5-yl)-benzenamine dihydrochloride; GYKI 53655: 1-(4-aminophenyl)-3-methylcarbamyl-4-methyl-3,4-dihydro-7,8-methylenedioxy-5H-2,3-benzodiazepine hydrochloride; MK 801:(5S,10R)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate or dizocilpine hydrogen maleate; NBQX: 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo[f]quinoxaline-7-sulfonamide; SYM 2081: (2S,4R)-4-methylglutamic acid; UBP 310: (S)-1-(2-amino-2-carboxyethyl)-3-(2-carboxy-thiophene-3-yl-methyl)-5-methylpyrimidine-2,4-dione selective antagonist of GluK1/GluK3. (b) Agonists of iGluRs. (c) Family of ionotropic glutamate receptor subunits.
Part (c) adapted from Wollmuth LP, Sobolevsky AI (2004) Structure and gating of the glutamate receptor ion channel. Trend. Neurosci.27, 321–328, with permission.
From Cellular and Molecular Neurophysiology, Fourth Edition.
Copyright © 2015 Elsevier Ltd. All rights reserved. 3

4. 4 Figure 10.1 Pharmacology of ionotropic glutamate receptors.(cont.)
(a) AMPA: α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid; ATPA : (RS)-2-amino-3-(3-hydroxy-5-tert-butylisoxazol-4-yl)propanoic acid selective agonist of GluK1-containing receptors; CNQX : 6-cyano- 7-nitroquinoxaline-2,3-dione; D-AP5 :DL-2-amino-5-phosphonopentanoic acid; DNQX: 6,7-dinitroquinoxaline-2,3-dione; GYKI 52466: 4-(8-methyl-9H-1,3-dioxolo[4,5-h,2,3]benzodiazepin-5-yl)-benzenamine dihydrochloride; GYKI 53655: 1-(4-aminophenyl)-3-methylcarbamyl-4-methyl-3,4-dihydro-7,8-methylenedioxy-5H-2,3-benzodiazepine hydrochloride; MK 801:(5S,10R)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate or dizocilpine hydrogen maleate; NBQX: 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo[f]quinoxaline-7-sulfonamide; SYM 2081: (2S,4R)-4-methylglutamic acid; UBP 310: (S)-1-(2-amino-2-carboxyethyl)-3-(2-carboxy-thiophene-3-yl-methyl)-5-methylpyrimidine-2,4-dione selective antagonist of GluK1/GluK3. (b) Agonists of iGluRs. (c) Family of ionotropic glutamate receptor subunits.
Part (c) adapted from Wollmuth LP, Sobolevsky AI (2004) Structure and gating of the glutamate receptor ion channel. Trend. Neurosci.27, 321–328, with permission.
From Cellular and Molecular Neurophysiology, Fourth Edition.
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5. 5 Figure 10.2 Organization of domains and subunits in iGluRs.
(a) See text. (b) iGluR subunits are composed of distinct domains including the amino terminal domain (ATD), ligand-binding domain (LBD), transmembrane domain (TMD), and carboxyl terminal domain (CTD). The ion channel is formed by the membrane-embedded domains 1, 2 (P-loop), 3 and 4. The iGluR S1S2 constructs are generated by deleting the ATD, coupling the end of S1 to the beginning of S2 via a Gly-Thr (GT) linker and deleting the final transmembrane segment by ending the polypeptide near the end of S2 (X). For GluA and GluK receptor subunits and for GluN2 subunit, the S1S2 complex forms the glutamate-binding site (agonist), whereas the GluN1 subunit forms the glycine-binding site. (c) Crystal structure of the homotetrameric full-length GluA2 receptors showing the pattern of subunit arrangement and domain organization in the tetrameric assembly. The four subunits are colored as blue, yellow, green and magenta.
Part (b) from Furukawa H (2012) Structure and function of glutamate receptor amino terminal domains. J. Physiol.590, 63–72, with permission. Part (c) from Sobolevsky AI, Rosconi MP, Gouaux E (2009) X-ray structure, symmetry and mechanism of an AMPA-subtype glutamate receptor. Nature462, 745–756, with permission.
From Cellular and Molecular Neurophysiology, Fourth Edition.
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6. 6 From Cellular and Molecular Neurophysiology, Fourth Edition.
Figure 10.3 iGluR-mediated EPSPs and their components.
(a) Current clamp recordings of a presynaptic glutamatergic pyramidal neuron and a postsynaptic interneuron in the cat neocortex in vitro. A spike triggered by a current pulse in the presynaptic neuron evokes a single-spike EPSP in the postsynaptic interneuron (intracellular recordings). (b) The same experiment in the hippocampus in vitro. In response to a stronger stimulation of the presynaptic pyramidal neuron and in the absence of external Mg2+, a postsynaptic EPSP of larger amplitude (control) with a fast rising phase and a long duration (sometimes up to 500 ms) is recorded. The same experiment in the presence of APV (APV, 33 μM). Picrotoxin is added to the extracellular solution in order to block GABAA synaptic receptors. (c) In response to the stimulation of afferent fibers, a control EPSP is recorded from a hippocampal interneuron. In the presence of blockers of NMDA (APV), AMPA (GYKI 53655), GABAB (CGP 55845) and GABAA (bicuculline) receptors, a low-amplitude component is still present. It is mediated by kainate receptors since it is totally blocked by LY
Part (a) from Buhl EH, Tamas G, Szilagyi T et al. (1997) Effect, number and location of synapses made by single pyramidal cells onto spiny interneurons of cat visual cortex. J. Physiol. (Lond.) 500, 689–713, with permission. Part (b) from Forsythe ID, Westbrook GL (1988) Slow excitatory postsynaptic currents mediated by N-methyl- d-aspartate receptors on cultured mouse central neurons. J. Physiol. (Lond.) 396, 515–533, with permission. Part (c) from Cossart R, Esclapez M, Hirsch J et al. (1998) GluR5 kainate receptor activation in interneurons increases tonic inhibition of pyramidal cells. Nat. Neurosci. 1, 470–478, with permission.
From Cellular and Molecular Neurophysiology, Fourth Edition.
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7. Figure 10.4 Electrophysiological properties of native AMPA receptor-channel.
(a) Patch clamp recording (outside-out configuration) of the activity of a quisqualate-activated channel. When the membrane is held at −60 mV, the unitary current iq is inward (downward deflection). At +60 mV or +80 mV, iq is outward (upward deflection). (b) Unitary current amplitude histogram (in pA). Currents recorded at the same voltage but from different membrane patches. (c)iq/V curve obtained from the averages of unitary currents recorded from a homogeneous population of channels (8 pS population). Intrapipette solution (in mM): 140 CsCI, 5 K-EGTA, 0.5 CaCl2; extracelluar solution: 140 Nacl, 2.8 KCl, 1 CaCl2.
Parts (a) and (c) from Ascher P, Nowak L (1988) Quisqualate and kainate-activated channels in mouse central neurons in culture. J. Physiol. (Lond.)399, 227–245, with permission. Part (b) from Cull-Candy SG, Usowicz MM (1987) Patch clamp recording from single glutamate-receptor channels. TIPS8, 218–224, with permission.
From Cellular and Molecular Neurophysiology, Fourth Edition.
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8. 8 Figure 10.5 Functional properties of AMPA receptors.
(a) Linear representation of the M2 segment (P-loop) of GluA receptors and site of editing (Q/R site) in this segment. (b) Comparison of whole-cell currents evoked by pulse application (25 s, bars) of a glutamate agonist to homomeric GluA1(Q) (left) or heteromeric GluA1(Q) + GluA2(R) (right) channels expressed in oocytes and recorded in normal Ringer (Na+) and Ca2+-Ringer (Ca2+) solutions. Oocytes were injected with a single GluA subunit cRNA (2 ng) or a combination of two types of GluA subunit cRNA (2 ng + 2 ng for 1:1 combination). Intrapipette solution (in mM): 250 CsCl, 250 CsF, 100 EGTA. Na+-external solution (in mM): 115 NaCl, 2.5 KCl, 1.8 CaCl2, 10 Hepes; Ca2+-external solution (in mM): 10 CaCl2, 10 Hepes. (c) I/V plots for whole-cell and outside-out patch responses to glutamate recorded from GluA2-lacking channels expressed in HEK cells. (left) Whole-cell responses are recorded on average 100 s after breakthrough, using a polyamine-free internal solution (red trace). Data from outside-out patches (black trace) are recorded using 60 μM spermine in the intrapipette solution; this concentration closely matches the complex I/V relationship of whole-cell responses over the range −100 to +100 mV. The two I/V plots are superimposed. (Right) I/V plot of the glutamate-induced current recorded in outside-out patches. The I/V plot is roughly linear.
Part (b) from Hollmann M, Hartley M, Heinemann S (1991) Ca2+ permeability of KA-AMPA-gated glutamate receptor channels depends on subunit composition. Science252, 851–853, with permission. Part (c) from Koh DS, Burnashev N, Jonas P (1995) Block of native Ca(2+)-permeable AMPA receptors in rat brain by intracellular polyamines generates double rectification. J. Physiol. 486, 305–312 and Bowie D, MayerML (1995) Inward rectification of both AMPA and kainate subtype glutamate receptors generated by polyamine-mediated ion channel block. Neuron15, 453–462.
From Cellular and Molecular Neurophysiology, Fourth Edition.
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9. 9 Figure 10.5 Functional properties of AMPA receptors.(cont.)
(a) Linear representation of the M2 segment (P-loop) of GluA receptors and site of editing (Q/R site) in this segment. (b) Comparison of whole-cell currents evoked by pulse application (25 s, bars) of a glutamate agonist to homomeric GluA1(Q) (left) or heteromeric GluA1(Q) + GluA2(R) (right) channels expressed in oocytes and recorded in normal Ringer (Na+) and Ca2+-Ringer (Ca2+) solutions. Oocytes were injected with a single GluA subunit cRNA (2 ng) or a combination of two types of GluA subunit cRNA (2 ng + 2 ng for 1:1 combination). Intrapipette solution (in mM): 250 CsCl, 250 CsF, 100 EGTA. Na+-external solution (in mM): 115 NaCl, 2.5 KCl, 1.8 CaCl2, 10 Hepes; Ca2+-external solution (in mM): 10 CaCl2, 10 Hepes. (c) I/V plots for whole-cell and outside-out patch responses to glutamate recorded from GluA2-lacking channels expressed in HEK cells. (left) Whole-cell responses are recorded on average 100 s after breakthrough, using a polyamine-free internal solution (red trace). Data from outside-out patches (black trace) are recorded using 60 μM spermine in the intrapipette solution; this concentration closely matches the complex I/V relationship of whole-cell responses over the range −100 to +100 mV. The two I/V plots are superimposed. (Right) I/V plot of the glutamate-induced current recorded in outside-out patches. The I/V plot is roughly linear.
Part (b) from Hollmann M, Hartley M, Heinemann S (1991) Ca2+ permeability of KA-AMPA-gated glutamate receptor channels depends on subunit composition. Science252, 851–853, with permission. Part (c) from Koh DS, Burnashev N, Jonas P (1995) Block of native Ca(2+)-permeable AMPA receptors in rat brain by intracellular polyamines generates double rectification. J. Physiol. 486, 305–312 and Bowie D, MayerML (1995) Inward rectification of both AMPA and kainate subtype glutamate receptors generated by polyamine-mediated ion channel block. Neuron15, 453–462.
From Cellular and Molecular Neurophysiology, Fourth Edition.
Copyright © 2015 Elsevier Ltd. All rights reserved. 9

10. 10 Figure 10.5 Functional properties of AMPA receptors.(cont.)
(a) Linear representation of the M2 segment (P-loop) of GluA receptors and site of editing (Q/R site) in this segment. (b) Comparison of whole-cell currents evoked by pulse application (25 s, bars) of a glutamate agonist to homomeric GluA1(Q) (left) or heteromeric GluA1(Q) + GluA2(R) (right) channels expressed in oocytes and recorded in normal Ringer (Na+) and Ca2+-Ringer (Ca2+) solutions. Oocytes were injected with a single GluA subunit cRNA (2 ng) or a combination of two types of GluA subunit cRNA (2 ng + 2 ng for 1:1 combination). Intrapipette solution (in mM): 250 CsCl, 250 CsF, 100 EGTA. Na+-external solution (in mM): 115 NaCl, 2.5 KCl, 1.8 CaCl2, 10 Hepes; Ca2+-external solution (in mM): 10 CaCl2, 10 Hepes. (c) I/V plots for whole-cell and outside-out patch responses to glutamate recorded from GluA2-lacking channels expressed in HEK cells. (left) Whole-cell responses are recorded on average 100 s after breakthrough, using a polyamine-free internal solution (red trace). Data from outside-out patches (black trace) are recorded using 60 μM spermine in the intrapipette solution; this concentration closely matches the complex I/V relationship of whole-cell responses over the range −100 to +100 mV. The two I/V plots are superimposed. (Right) I/V plot of the glutamate-induced current recorded in outside-out patches. The I/V plot is roughly linear.
Part (b) from Hollmann M, Hartley M, Heinemann S (1991) Ca2+ permeability of KA-AMPA-gated glutamate receptor channels depends on subunit composition. Science252, 851–853, with permission. Part (c) from Koh DS, Burnashev N, Jonas P (1995) Block of native Ca(2+)-permeable AMPA receptors in rat brain by intracellular polyamines generates double rectification. J. Physiol. 486, 305–312 and Bowie D, MayerML (1995) Inward rectification of both AMPA and kainate subtype glutamate receptors generated by polyamine-mediated ion channel block. Neuron15, 453–462.
From Cellular and Molecular Neurophysiology, Fourth Edition.
Copyright © 2015 Elsevier Ltd. All rights reserved. 10

11. Figure 10.6 Whole-cell kainate current in cerebellar granule cells.
(a) Whole-cell current (Ikai) evoked by the application of 10 μm of kainate in the presence of GYKI (100 μM) in a concanavalin A-treated granule cell. (b)Ikai/V relationship. Ikai is measured during 500 ms voltage ramps from −80 to +80 mV.
From Pemberton KE, Belcher SM, Ripellino JA, Howe JR (1998) High affinity kainate-type ion channels in rat cerebellar granule cells. J. Physiol. (Lond.)510, 401–420, with permission.
From Cellular and Molecular Neurophysiology, Fourth Edition.
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12. Figure 10.7 Correlation of functional properties of native kainate receptors and RNA editing of the Q/R site of the GluR6. (a) KAR subunits (GluK1–5) and splice variants. Black boxes represent membrane domains (M1–M4). Triangles depict sites of RNA editing, including the ‘Q/R’ site within both GluK1 and GluK2, which controls ion permeability of the channel. (b) Expressing homomeric GluK2(Q) and (c) expressing homomeric GluK2(R). Kainate (300 μM) is rapidly applied while holding the membrane potential at different voltages, from −70 to +50 mV. Insets show the current traces at these different voltages.
Part (a) from Contractor A, Mulle C, Swanson GT (2011) Kainate receptors coming of age: milestones of two decades of research. Trends Neurosci. 34, 154–163, with permission. Part (b) from Ruano D, Lambolez B, Rossier J et al. (1995) Kainate receptor subunits expressed in single cultured hippocampal neurons: molecular and functional variants by RNA editing. Neuron14, 1009–1017, with permission.
From Cellular and Molecular Neurophysiology, Fourth Edition.
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13. Figure 10.7 Correlation of functional properties of native kainate receptors and RNA editing of the Q/R site of the GluR6.(cont.) (a) KAR subunits (GluK1–5) and splice variants. Black boxes represent membrane domains (M1–M4). Triangles depict sites of RNA editing, including the ‘Q/R’ site within both GluK1 and GluK2, which controls ion permeability of the channel. (b) Expressing homomeric GluK2(Q) and (c) expressing homomeric GluK2(R). Kainate (300 μM) is rapidly applied while holding the membrane potential at different voltages, from −70 to +50 mV. Insets show the current traces at these different voltages.
Part (a) from Contractor A, Mulle C, Swanson GT (2011) Kainate receptors coming of age: milestones of two decades of research. Trends Neurosci. 34, 154–163, with permission. Part (b) from Ruano D, Lambolez B, Rossier J et al. (1995) Kainate receptor subunits expressed in single cultured hippocampal neurons: molecular and functional variants by RNA editing. Neuron14, 1009–1017, with permission.
From Cellular and Molecular Neurophysiology, Fourth Edition.
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14. Figure 10.8 Unitary NMDA current in the absence of extracellular Mg2+.
(a) Outside-out patch clamp recordings of the activity of an NMDA (10 μM) activated channel at two holding potentials, −60 and +40 mV. (b)iN/V relation obtained from the averages of unitary currents iN recorded from a homogeneous population of channels (a population that shows a 40–50 pS unitary conductance). Intrapipette solution (in mM): 140 CsCl, 5K-EGTA, 0.5 CaCl2; extracellular solution: 140 NaCl, 2.8 KCl, 1 CaCl2.
Part (a) from Ascher P, Bergestovski P, Nowak L (1988) N-methyl- d-aspartate-activated channels of mouse central neurons in magnesium free solutions. J. Physiol. (Lond.)399, 207–226, with permission. Part (b) from Cull-Candy SG, Usowicz MM (1987) Patch clamp recording from single glutamate-receptor channels. TIPS8, 218–224, with permission.
From Cellular and Molecular Neurophysiology, Fourth Edition.
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15. Figure 10.9 Optical measurements of intracellular Ca2+ concentration changes during an NMDA-evoked response.
The Ca2+-sensitive dye Arsenazo III is used. The absorption coefficient of this dye varies at certain wavelengths when it complexes Ca2+ ions. The activity of cultured spinal neurons is recorded in the whole-cell patch clamp configuration in the absence of external Mg2+. (a) Pressure application of 1 μ M of NMDA (20 ms) in the presence of 2.5 m M of Ca2+ in the extracellular milieu evokes an inward current IN (top trace). During this response there is an increase of [Ca2+]l (bottom trace). (b) Reversal potential of the whole-cell NMDA current (IN) as a function of extracellular [Ca2+]o. Currents activated by the application of 1 m M of NMDA are recorded at different membrane potentials in the presence of 1 m M (left) or 20 m M (right) of [Ca2+]o.
Part (a) from Mayer ML, MacDermott AB, Westbrook GL et al. (1987) Agonist- and voltage-gated calcium entry in cultured mouse spinal chord neurons under voltage clamp using Arsenazo III. J. Neurosci.7, 3230–3244, with permission. Part (b) from MacDermott AB, Mayer ML, Westbrook GL et al. (1986) NMDA-receptor activation increases cytoplasmic calcium concentration in cultured spinal neurons. Nature321, 519–522, with permission.
From Cellular and Molecular Neurophysiology, Fourth Edition.
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16. Figure 10.10 NMDA channel block by extracellular Mg2+ ions.
(a) Outside-out patch clamp recording of the activity of cultured central neurons. Application of NMDA (10 μM) in the absence of external Mg2+ ions (O) and in the presence of Mg2+ (10, 50, 100 μM) at VH = −60 mV. At VH = +40 mV, iN is outward. (b) Voltage sensitivity of the NMDA response in the presence of extracellular Mg2+ ions. The total current IN is recorded in the whole-cell patch clamp configuration in the absence (o) and in the presence (■) of 500 μM of Mg2+.
Part (a) from Ascher P, Nowak L (1988) The role of divalent cations in the N-methyl- d-aspartate responses of the mouse central neurons in culture. J. Physiol. (Lond.)399, 247–266, with permission. Part (b) from Nowak L, Bregestovski P, Ascher P et al. (1984) Magnesium free glutamate-activated channels in mouse central neurones. Nature307, 463–465, with permission. C – Closed state; O – Open state.
From Cellular and Molecular Neurophysiology, Fourth Edition.
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17. 17 From Cellular and Molecular Neurophysiology, Fourth Edition.
Figure Permeability of NMDA channels to divalent cations.
(a, top) Seven NMDA receptor (NMDAR) subunits have been identified: GluN1, GluN2A–GluN2D and GluN3A and GluN3B. Subunit heterogeneity is further enhanced by alternative splicing of GluN1 and GluN3A subunits. M1–M4 indicate membrane segments. (a, bottom) the GluN subunits carry an asparagine residue (N) in the M2 segment in a position homologous to the Q/R site of AMPAR and KAR. The GluN2 subunits also carry an asparagine residue at site N + 1. (b,c) Reduction of Ca2+ permeability and channel block by extracellular Mg2+ in a mutant channel where asparagine (N) in the M2 segment of the NR1 subunit is replaced by arginine (R). (b) Whole-cell current elicited by 100 μM of glutamate (bar) at VH = −60 mV in high Na+ (inward current) or high Ca2+ (small outward current) extracellular solution. (c) Whole-cell IN/V relations in (1) divalent ion-free external solution and (2) after adding 0.5 mM of Mg2+ to the external solution. (d,e) Difference in channel block by extracellular Mg2+ between wild-type and mutant NMDA receptor-channels. In the GluN2A (N595Q) subunit, one asparagine (N) in the M2 segment is replaced by glutamine (Q) by site-directed mutagenesis. The whole-cell current evoked by glutamate is recorded in (1) divalent ion-free Ringer and (2) after addition of 0.1 mM of Mg2+ as a function of membrane potential from (d) wild-type channels and (e) a mutant channel. Extracellular high Na+ solution (in mM): 140 Nacl, 5 HEPES: high Ca2+ solution: 110 CaCl2, 5 HEPES. Divalent ion-free Ringer’s solution: 135 NaCl, 5.4 KCl, 5 HEPES. (f) Amino acid residue sequence alignment of the M3 segment of GluN2A–D subunits. The asterisk marks the GluN2 S/L site.
Part (a, top) Adapted from Paoletti P, Bellone C, Zhou Q (2013) NMDA receptor subunit diversity: impact on receptor properties, synaptic plasticity and disease. Nat. Rev. Neurosci.14, 383–400, with permission. Part (a, bottom) from Wisden W, Seeburg PH (1993) Mammalian ionotropic glutamate receptors. Curr. Opin. Neurobiol.3, 291–298, with permission. Parts (b)–(e) from Burnashev N, Schoepfer R, Monyer H et al. (1992) Control of calcium permeability and magnesium blockade in the NMDA receptor. Science257, 1415–1419, with permission. Part (f) from Siegler Retchless B, Gao W, Johnson JW (2012) A single GluN2 subunit residue controls NMDA receptor channel properties via intersubunit interaction. Nat. Neurosci. 15, 406–413, with permission.
From Cellular and Molecular Neurophysiology, Fourth Edition.
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18. 18 From Cellular and Molecular Neurophysiology, Fourth Edition.
Figure Permeability of NMDA channels to divalent cations.(cont.)
(a, top) Seven NMDA receptor (NMDAR) subunits have been identified: GluN1, GluN2A–GluN2D and GluN3A and GluN3B. Subunit heterogeneity is further enhanced by alternative splicing of GluN1 and GluN3A subunits. M1–M4 indicate membrane segments. (a, bottom) the GluN subunits carry an asparagine residue (N) in the M2 segment in a position homologous to the Q/R site of AMPAR and KAR. The GluN2 subunits also carry an asparagine residue at site N + 1. (b,c) Reduction of Ca2+ permeability and channel block by extracellular Mg2+ in a mutant channel where asparagine (N) in the M2 segment of the NR1 subunit is replaced by arginine (R). (b) Whole-cell current elicited by 100 μM of glutamate (bar) at VH = −60 mV in high Na+ (inward current) or high Ca2+ (small outward current) extracellular solution. (c) Whole-cell IN/V relations in (1) divalent ion-free external solution and (2) after adding 0.5 mM of Mg2+ to the external solution. (d,e) Difference in channel block by extracellular Mg2+ between wild-type and mutant NMDA receptor-channels. In the GluN2A (N595Q) subunit, one asparagine (N) in the M2 segment is replaced by glutamine (Q) by site-directed mutagenesis. The whole-cell current evoked by glutamate is recorded in (1) divalent ion-free Ringer and (2) after addition of 0.1 mM of Mg2+ as a function of membrane potential from (d) wild-type channels and (e) a mutant channel. Extracellular high Na+ solution (in mM): 140 Nacl, 5 HEPES: high Ca2+ solution: 110 CaCl2, 5 HEPES. Divalent ion-free Ringer’s solution: 135 NaCl, 5.4 KCl, 5 HEPES. (f) Amino acid residue sequence alignment of the M3 segment of GluN2A–D subunits. The asterisk marks the GluN2 S/L site.
Part (a, top) Adapted from Paoletti P, Bellone C, Zhou Q (2013) NMDA receptor subunit diversity: impact on receptor properties, synaptic plasticity and disease. Nat. Rev. Neurosci.14, 383–400, with permission. Part (a, bottom) from Wisden W, Seeburg PH (1993) Mammalian ionotropic glutamate receptors. Curr. Opin. Neurobiol.3, 291–298, with permission. Parts (b)–(e) from Burnashev N, Schoepfer R, Monyer H et al. (1992) Control of calcium permeability and magnesium blockade in the NMDA receptor. Science257, 1415–1419, with permission. Part (f) from Siegler Retchless B, Gao W, Johnson JW (2012) A single GluN2 subunit residue controls NMDA receptor channel properties via intersubunit interaction. Nat. Neurosci. 15, 406–413, with permission.
From Cellular and Molecular Neurophysiology, Fourth Edition.
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19. 19 Figure 10.12 Potentiation of the NMDA response by glycine.
(a, top) Whole-cell currents evoked in cultured central neurons in response to 10 μM of NMDA or 10 μM of glutamate at VH = −50 mV in the absence or presence of 1 μM of glycine (Gly). (a, bottom) The same experiment with quisqualate (Quis) or kainate (Kai) applications. Glycine by itself does not trigger an inward current at any concentration, through either NMDA or non-NMDA channels. (b) Whole-cell inward current (66 ± 13 nA) evoked in Xenopus oocytes which express NMDA receptors, in response to 300 μM of NMDA at VH = −60 mV in the absence of presence of 3 μM of glycine. In (a) and (b) the extracellular solution is devoid of Mg2+ ions.
Part (a) from Johnson JW, Ascher P (1987) Glycine potentiates the NMDA response in cultured mouse brain neurons. Nature325, 529–531, with permission. Part (b) from Kleckner N, Dingledine R (1988) Requirements for glycine in activation of NMDA receptors expressed in Xenopus oocytes. Science241, 835–837, with permission.
From Cellular and Molecular Neurophysiology, Fourth Edition.
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20. Figure 10.13 The glutamatergic synapse.
Functional scheme of a glutamatergic synapse where ionotropic and metabotropic glutamate receptors are co-localized. Presynaptic receptors are omitted. The enzymes (1 to 3) and mitochondria are carried to axon terminals via anterograde axonal transports. Glutamate synthesized in mitochondria of the presynaptic element is transported actively into synaptic vesicles by a vesicular carrier. A percentage of the glutamate released in the synaptic cleft is uptaken into presynaptic terminals and glial cells by transporters. Inset shows iGluRs antagonists.
Inset from Mody I (1998) Interneurons and the ghost of the sea. Nat. Neurosci.1, 434–436, with permission.
From Cellular and Molecular Neurophysiology, Fourth Edition.
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21. Figure Postsynaptic inward current evoked by the stimulation of a glutamatergic presynaptic neuron.
(a) Whole-cell postsynaptic inward current (EPSC) recorded VH = −46 mV in the absence of Mg2+, in response to the activation of a presynaptic glutamatergic neuron. The peak current decays with a time constant τ1 = 4.2 ms and the slow components decays with a time constant τ2 = 81.8 ms. (b) The EPSC is recorded at different VH in the presence of Mg2+ (100 μM). (c) The EPSC is recorded at different VH in the presence of 33 μM of D-APV and in the absence of extracellular Mg2+. Picrotoxin (10–100 μM) is added to the extracellular solution to block GABAergic inhibitory synaptic activity.
From Forsythe ID, Westbrook GL (1988) Slow excitatory postsynaptic currents mediated by N-methyl- d-aspartate receptors on cultured mouse central neurons. J. Physiol. (Lond.)396, 515–533, with permission.
From Cellular and Molecular Neurophysiology, Fourth Edition.
Copyright © 2015 Elsevier Ltd. All rights reserved. 21

22. Figure 10.15 The AMPA- and kainate-mediated component of EPSCs.
Experiments performed in slices of the rat hippocampus (CA1 region). (a) Averaged EPSCs recorded from an interneuron (I) in control conditions (continuous presence of 100 μM of D-APV), after bath application of 70 μM of GYKI and after addition of 100 μM of CNQX. Middle traces are the same EPSCs at high gain. (b) The same experiment performed in pyramidal cells (pyR). VH in (a) and (b) is −80 mV.
From Frerking M, Malenka RC, Nicoll RA (1998) Synaptic activation of kainate receptors on hippocampal interneurons. Nat. Neurosci.1, 479–486, with permission.
From Cellular and Molecular Neurophysiology, Fourth Edition.
Copyright © 2015 Elsevier Ltd. All rights reserved. 22