4. Κανάλια ιόντων και μεταγωγή σήματος

1.

2.

3. - - - -

4. - (ligand-gated ion channel receptors)

Cys-loop

Gaba : NMDA

5. G-

6. (cGMP, cAMP) (3)

4o

80

1.

, . , , .. .

, .

. . - , .

-- . , , - . , , , GABA, , , , ( 4.1). . , - Ca2+ Ca2+ . Ca2+ .

4.1 . Kraus, Biochemistry of Signal Transduction, 2000.

81

( 4.2). - (ligand gated ion channels), G- -. , -.

- . -. : - Na+/K+ - Cl-. , .

4.2 . ,

. Ca2+ . , , . .

(), , G (), . G , cAMP. cAMP cAMP-, . Eric Kandel, , 2000.

82

2. : -

Na+ Cl-

, K+ , . , , . , , :

1. Na+ K+ . - Na+/K+.

2. K+ K+.

, . , 30mV 50 mV . , : Na+, , , , , K+ Cl- , . , , . : - Na+ , (msec) , ( +30mV). , K+, K+, , . msec, ( 4.3).

, - Ca2+ ( 4.4).

140 mM K+ 4 mM K+ 12 mM Na+ 150 mM Na+ 4 mM Cl- 120 mM Cl- 148 mM A- ( ) 34 mM A-

83

4.3 . 1. . 2. a+

. 3. . 4. + . 5. .

4.4 . , .

1.

2.

3. 4.

5.

84

3. -

. :

- , .

-, , .

(effectors) . , Gai , G- (GPCRs).

, , . NMDA .

, , , .

-

- . , , a+, + Ca2+. : , 0,5 2 msec, a+, + Ca2+. msec. , 2 msec . . , , . , . : , . . . .

I: a+, + Ca2+ . , a+ 12 .

85

Ca2+ Ca2+ 1000 .

-

- 140 , (review Yo et al, 2005).

- a+ Eliktrophorus electricus, Ca2+ + shaker Drosophila. a+ Ca2+ , + .

- Na+ Ca2+ -, 1-4 . - 4 (-IV).

4.5 . - - Na+ ( Ca2+). - Na+ Ca2+ 4 (-IV). Yu et al, Overview of molecular relationships in the voltage-gated ion channel superfamily, Pharmacological Reviews, 2005, 57, 387-395. . - - (auxiliary subunit) Na+.

.

.

86

- + -, (-IV) - Na+ Ca2+.

4.6 - + . -

+ 4 -, I-IV Na+ Ca2+. - 6 (S1-S6),

-. S4 , SS1 SS2 . .

. 4.7 - -. 6 (S1-S6), -. S4 , SS1 SS2 .

, . , , , / . (gating current). , .

87

S4 . S4 . 4. 8 S4. S4 + . S4. (R1, R2, R3, R4, K5, R6).

. . . S4 , , . S4 - - . .

4.9

( S4), , .

88

- (-IV). , , , , . a+ , , . SS1 SS2 ( 4.7). SS1 SS2, . , , SS1 SS2 . , S6 .

-

, . , msec sec, . a+, , , . a+: Na+

- ,

- . - 200 . , , .

, .

Ion passage and ion selectivity

4.10 Na+,

.

89

. .

-

- - , , (. ) , . , .

Na+ (1 4). , - , C- . , . Cys .

Ca2+ 4 (2, , , ). 2 , ( 2 ) . - . 4 , , .

+ -, - . .

4.11 - . Yu et al,

Overview of molecular relationships in the voltage-gated ion channel superfamily, Pharmacological Reviews, 2005, 57, 387-395.

90

4.12 - Ca2+. 1 2, , .

4. - (Ligand-gated ion channel receptors)

- - .

3 :

(2), , , - C- . 7 2, . 22/3, 21/5, 22/6, 24/6.

cys-loop , -, 5 (2) . 4 , C- - . , / / . , . 3 4 , . Cys-loop ( nAChR, 5-3, C-Zinc activated ion channel Zn2+) (- GABAA, GABAC ).

, 4 . - , , P loop, . C- . NMDA ( NR1 R2A, NR2B, NR2C, NR2D), AMPA .

91

4.13 (A) :

(2), (GluR) Cys-loop (nAChR, 5HT3, ZAC, GABAA gly). (B) 4 /, . . Hogg RC., Buisson B., Bertrand D., Allosteric modulation of ligand-gated channels, Biochemical Pharmacology, 2005, 70, 1267-1276.

- , , : a+ Ca2+ , 5-3 , Cl- GABAA, GABAC . (depolarization) -. , 5-3 ( NMDA, AMPA ), . (hyperpolarization) -. GABAA GABAC , .

92

, msec, (fast responding receptors), G, , G, .

- Cys-loop

O 5 (subunits) - : 4 , - , - .

4 EN (-V). 20-30

-, . ( 20) .

4.14 (A) GABAA. 5 (22)

. : 4 , NH2 COOH , 2 . Bormann J., The ABC of GABA receptors. TiPS, 2000, 21, 16-19. (B) 4 22 GABAA. 4 (1, 2, 3, 4). G.B. Smith and R.W.Olsen, TiPS 1995, 16, 162-167.

( ,

) ( 5,6 ). , . (Pro) .

Cys-

93

, (Ser248, Ser254, Ser256, Ser262) , (non competitive blockers)* ( 4.15, 4.16). (. , , ) (), , N- -.

4.15 . -. 4 (1-4), 20-30 - . . . 2 . H.R. Arias, Brain Res.Rev., 1997, 25, 133-191.

() , 2 ( ) . CPZ (), (-), TID Arias H.R., Brain Res. Review, 1997, 25, 133-191.

* , . (D). . (J-P. Changeux et al, Achetylcholine receptor: an allosteric protein, Science, 1984, 225, 1335-1344).

A B

94

4.16 .

-. (high affinity site) , 30 , (low affinity sites) . J-P. Changeux et al, Science, 1984, 225, 1335-1343. 1 E 2-

3 .

: , GABAA, Cl-, , , Na+ Ca2+. - , (driving force) .

2 . -

, cAMP. .

AMINOTEIKO AO (NH2-TERMINAL). , - , (Cys139 Cys153), S-S. Cys-Cys ,

, . . , , , ( - -). (2).

. (2) GABA (22) 5-3, -, . . , . 2001, Changeux, ( ACh). ( ) -, , ACh - - ( 4.17).

95

ACh

ACh

ACh

4.17 () . ()

. () , . . . () , ACh, - -. () , Galzi and Changeux, 1995. -. () , Changeux 2001. -, - -. Lester et al, Cys-loop receptors: New twists and turns, Trends in Neurosciences, 2004, 27, 329-336. - - GABA

. GABA . GABA .

(COOH-TERMINAL).

G, , - .

96

. , . , . , . . 2-3 nm .

4.18 , . ) . , . . . ) 2 -, . Unwin, 1993.

.

-

97

.

prefilter

M2M2M2

4.19 () 2 .

( ) . .

() 2 . . 2 . (): 2 . . (): 2 . 2 . ( ) , . Unwin, 1995. , 2 . . , . , - , , 2. 2 . , . 2 , . .

98

4.20 ACh, ACh. ACh Torpedo , Unwin et al (2003). (2). Ch () (). 2 - (), () . Colquhoun D., and Sivilotti L., Function and structure in glycine receptors and some of their relatives, Trends in Neurosciences, 2004, 27, 327-344.

4.21 ACh, ACh . Ch . Lester H., Dibas M., Dahan D., Leite JF., Dougherty DA., Cys-loop receptors: New twists and turns, Trends in Neurosiences, 2004, 27, 329-336.

GABA :

GABA . GABA ( , 17-20% GABA, ~6.000 ),

, -

, -

99

- GABAA, Cl- .

GABA :

GABA ,

GABA . GABA , 2, -

..

(.

). () GABA, , GABA. 6 - GABA ,

GABA.

E 4.22 GABA. GABA GABA

. GABAA - Cl-, () , (GABA spillover) GABAA , () .

Semyanov A, Walker M., Kullman D., Silver R., Tonically active GABAA receptors: modulating gain and maintaining the tone, Trends in Neurosciences 2004, 27, 262-269.

GABA :

(. CA1 ). () GABA, , GABA. 6 - GABA , GABA.

GABA , GABA . GABA ,

100

2, - . , GABA GABA . GABA (ipsc) , GABA.

GABA

1. GABA: GABA -

: (1-6), (1-3), (1-3), , , . , . , GABA , .

GABA , . , .

(ipsps) GABA. ,

101

. , CA1 GABA 1, 2 5 . , ipsps . ; , .

2. - GABA

, GABA . .

6 ( ) . 6 .

3. GABA,

. GABA , . , GABA , GA , .

4. GABA () () ,

GABA . , .

5. Cl- Cl- GABA

. , K+/ Cl- Cl-, Cl- GABA.

6. GABA

, . , GABA. /. Ser410 2 (Akt), GABA . , 2 / 3 .

7. GABA

. GABA , , , 5-, ACh .

102

GABA GABA. GABA , GABA GABA . GPCR , GABA, Ca2+.

4.23 GABAA. , : (1) . (2) . (3) , C (PKC). (4) Cl-, K+/ Cl- (KCC2) . , : (5) Ca2+ (6) GABAB , (7) , , (8) , GB1 , (9) , , 5-, ACh. , , , (10). Mody I., Pearce R., Diversity of inhibitory neurotransmission trough GABAA receptors, Trends in Neurosciences, 2004, 27, 569-575.

-

- (Glu), . , , NMDA (--D-), AMPA (--3--5--4- ).

103

, NMDA . NMDA LTP ( ). , Ca2+. NMDA . NMDA :

.

AMPA NMDA. NMDA Mg2+ ( Zn2+),

. , , Na+.

Mg2+ NMDA.

NMDA a+ Ca2+.

4.24 NMDA, ,

( ) . . NMDA ( Mg2+ ) ( Na+). . , NMDA, Mg2+ , a+ Ca2+. Ca2+ .

104

NMDA

NMDA -, 4 : NR1 2 NR2A, NR2B, NR2C NR2D ( NR1-NR1-NR2A-NR2A).

3 (1, 3 4). 2 2 . 2 , , S1, S2 ( 4 , , ). COO- , .

4.25 NMDA. (NR1-NR4). 3 (1, 3 4). 2 2 .

4.26 . D (Amino Terminal Domain) 2 , . S1 , 1 , 2 3 . 3 4 S2 . S1 S2 . CTD (Carboxy Terminal Domain) . Wollmuth L., Sobolevsky A., Structure and gating of the glutamate receptor ion channel, Trends in eurosciences, 2004, 27, 321-328.

105

2 . 2 -, , 1 3 . Q/R/N . (Q), (R) , () DA ( 4.28).

4.27 2 .

+ + . + 6 ( 4.6). , + . 2 +. . .

4.28 +. + . 2 ( ) & Q/R/N . Wollmuth L., Sobolevsky A., Structure and gating of the glutamate receptor ion channel, Trends in eurosciences, 2004, 27, 321-328.

2

3

106

NMDA 2 NMDA . QRN . 4.29 -615 () R2 616 (+1) R1. . NMDA Ca2+ Mg2+. , NMDA Mg2+.

4.29 NMDA Mg2+. (616)

2 R1 (615) NR2B . Ca2+ Mg2+.

NMDA

Q/R/N , 3 -. SYTAANLAAF, . NMDA SYTAANLAAF. 4.30, SYTAANLAAF 3 . , - 3 . AMPA . NMDA Ca2+ AMPA. - 3. S1 S2 S2 8 Amstrong ( ). 3, 3 . 3

107

2 , ( 4.31).

4.30 () 2 3 - . (Q582) Q/R/N 2 - SYTAANLAAF ( ) 3 -. 3 , . () SYTAANLAAF 3 , DA ( ) ( ). Wollmuth L., Sobolevsky A., Structure and gating of the glutamate receptor ion channel, Trends in eurosciences, 2004, 27, 321-328.

4.31 .

.

.

3

R1 R2

108

NMDA, -, . NMDA , PCP, .

4.32 NMDA.

5. G-

G-. , , G-. GPCR G-. . + Gi Gq. GIRK ( Kir) (G protein-coupled inwardly rectifying potassium channels: G- +) , , . GPCRs ( 2, 2, D2, 5-1, GABA), G . GIRK, +, + ( ). , GIRK ACh, 2 .

109

GPCR, - G, GDP GTP. G-GTP G-, G . G GIRK G . Gi, G GIRK . + ( +, + ). G GTP GDP, G . Gq, GIRK , . Gq PLC, (2), (3) (DAG). PKC, , .

4.33 GIRK, G . . Gi , G , . . Gq , PKC 2 . Breitwieser GE, GIRK channels: hierarchy of control. Focus on "PKC-delta sensitizes Kir3.1/3.2 channels to changes in membrane phospholipid levels after M3 receptor activation in HEK-293 cells". Am J Physiol Cell Physiol. 2005, 289, C509-11.

+,

. + , . , +, , ( ) Mg2+, .

110

GIRK . 4 . (1 2) . .

4.34 G (GIRK).

(1 2) , . .

6.

(CAMP, cGMP) (3).

, cGMP cAMP, (cGMP- ) (cAMP- ) . : (). , G- (), - (effector). cGMP. cGMP. cGMP Ca2+ Na+ . cGMP . cGMP , Ca2+ Na+ .

GIRKGIRKVoltagegatedK+Channel

111

4.35 . (R) . cis/trans . (R*) G- . Gt,a cGMP (PDE), cGMP GMP. PDE , , . PDE cGMP. cGMP Na+ Ca2+ . cGMP , Na+ Ca2+ . . , cGMP cGMP . cGMP , Na+ Ca2+ .

: GPCR. , G- , cAMP. cAMP Ca2+ Na+ . cAMP , Ca2+ Na+ , .

112

4.36 . (R) . , G- (AC), cAMP. cAMP Ca2+ Na+. cAMP , Ca2+ Na+ , . Trudeau M., Zagotta W., Calcium/calmodulin modulation of olfactory and rod cyclic nucleotide-gated ion channels, J. Biol. Chem., 2003, 278, 21, 18705-18708.

cGMP cAMP . 4 . 6 , - - . +,

. cAMP/cGMP C- - (, , C) 8 (1-8).

4.37 ( 4) . cAMP/cGMP - (, , C) 8 (1-8). Biel M et al, Arch. Pharmacol., 1998, 358.

(CNG: cyclic

nucleotide gated ion channels) Ca2+. Ca2+ , Ca2+/, -

113

(cAMP, cGMP). feed-back -. 4.38 (CNG) (). cAMP/cGMP , Ca2+/ - . Ca2+/ - C- , cAMP/cGMP. Trudeau M., Zagotta W., Calcium/calmodulin modulation of olfactory and rod cyclic nucleotide-gated ion channels, J. Biol. Chem., 2003, 278, 21, 18705-18708.

(3)

InsP3, Ca2+ (InsP3). 3 Ca2+ . 15.000 , , Ca2+ .

- 3

1,4,5 (IP3) Ca2+ Ca2+ IP3. , , . IP3Rs : Ca2+ , Golgi .

114

4.39 () P3. 6 2 , NH2- IP3. 5 6 . () 4 IP3R. (,) . Bosanac et al, Structural insights into the regulatory mechanisms of IP3 receptors, Biochimica et Biophysica Acta 2004, 1742, 89-102. IP3Rs . ( 2700 ) (R) , - C- . 5 6, IP3Rs.

IP3R, - 226-576, IP3. IP3Rs 4 . 3 3 - .

3 1 TMR (~1600 ) , IP3Rs. , , IP3Rs : 3, .

IP3Rs 3 Ca2+. 3 - IP3R Ca2+ ( IP3) Ca2+. 3 Ca2+

()

() () ()

115

- C- , .

4.40 - P3. : , Ca2+, 3 ( ). Taylor C., da Fonseca C., Morris E., IP3 receptors: he search for structure, Trends in Biological Sciences, 2004, 29, 210-219.

4.41 - 3. ()

, (suppressor) - 1 (gatekeeper) C- 2 , . () 3 - , Ca2+. 3 Ca2+ - C- , . () Ca2+- - (gatekeeper) C- . Taylor C., et al, TiPs, 2004, 29, 210-219.

-

() () ()

116

Ca2+ . 3 20 Ca2+. (CAM: calmodulin), -, Ca2+/- (CAM-kinases).

: , G ( G ) cAMP, cGMP, 3.

1. GPCR, G

2. C

IP3 DAG 3. (DAG)

4. IP3 - Ca2+

5. To Ca2+

6. Ca2+ , -.

117

-

GPCR - G -

GPCR G Effector -

4.42 , . ) -: . ) G: . ) (cAMP, cGMP), .

Arias H.R., Topology of ligand binding sites on the nicotinic acetylcholine receptor, Brain Res. Rev., 1997, 25, 133-191.

Arias H.R., Localization of agonist and competitive antagonist binding sites on nicotinic acetylcholine receptors, Neurochem Int. 2000, 36, 595-645.

Barnard E.A., The transmitter-gated channels: a range of receptor types and structures, TiPS, 1996, 17, 305-308

Biel M., Sautter A., Ludwig A, Hoffmann F., Zong X., Cyclic nucleotide-gated channels mediators of NO: cGMP-regulated processes, Naunyn-Schiedebergs Arch. Pharmacol., 1998, 358, 140-144.

Bradley J., Reisert J., Frings S., Regulation of cyclic nucleotide-gated channels, Current Opinion in Neurobiology, 2005, 15, 343-349.

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Cascio M., Structure and Function of the glycine receptorand related nicotinicoid receptors, J. Biol. Chem., 2004, 279, 19383-19386.

()

()

()

118

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119

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