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Effect of 6-hydroxydopamine on dark adaptation in the rat retina
Pergamon Press
Lüe Sciences Vol . 13, pp . 15-21, 1973 . Printed in Great Britain
EFFECT OF 6-HYIROâ7mOPAMINB ON DARR ADAPTATION IN Tf~ RAT RETINA Schrs F. Pong and Lexie T. Graham, Jr . 9eotion of Neurobiology, The Institute of Peychiatrio Reaearoh~ Indiana University Medioal Center, Indianapolis, Indiana 46202 (Received 19 February 1973 ; in final form 9 Aprü 1973) SUl4~ARY 6-liydrozydopamine administered intravitreouaiy to Lightadapted rate appears to prevent the reocvery of retinal sensitivity during subsequent dark adaptation through its inter action xith the photochemical meohanisma . Adreaergio neurone, mainly containing dopamine, have been localised at the junction of the retinal inner nuclear and inner pleziform layers of all species so far investigated by the fluo~reaoenoe technique (1-5) .
Neuro
phyaiological studies have shown that ezogenously applied dopamine can inhibit the firing of retinal ganglion cells (6,7) . The ensymes responsible for the eetabolio degradation of aateoholaminea (8,9) and the uptabe and release of dopamine from the cat ratios follaring appropriate light stimulation have been demonstrated (10) . Thus, the inhibitory neurotransmitter role of retinal dopamine is strongly implicated .
However, littù direct information con-
cerning the functional role of dopamine in the retina ie available .
Attempts
to correlate the dopamine content, Wonoamine o~xidaae and aateohol-0 ..eethyl traneferaae activities to the state of adaptation have been reported, but not xithout some conflicting results (8,11,12) .
It is of interest to ants that
anitwls treated xith raserpine, a cateoholamioe-depleting agent, ezEsibit photophobia xhioh eight bs due to a blockade of aminergio inhibitory meohaniaes (1) . It has been xell established that 6-hydroxydopamine (6-HD) causes a selective and long-lasting daganeration of adrsnergio nerve tersinala and has been xidely employed recently as a phara~acologioal tool to study the fractions 15
18
6-Hydroxydopamine and Dark Adaptation
of adreoergio neurons (13) .
Vol . 13, No . 1
Due to its remarkable ohemical eympatheotaoy
property and to the feasibility of detecting changes provoked by pharmacological manipulation of retinal neurone by monitoring the eleotroretinogram (ERG) (1lt-16), and because of our interest in functional correlatione of neurotransmitters in retina, the effect of 6-HD on the ERG threshold of the rat retina was studied, Ptethode The appearanoe of the brave of the ERG xae used as a measure of retinal sensitivity and xae determined under experimental conditions similar to that used by Cone (17) as described previously (18),
The threshold ie the mini~nuo
Light stimulus neoeeeary to evoke a just-peroeptib]s response in the BR,G . Depending on the noise of the partioular preparation, the response aeleoted xae a potential of 10 to 20 pvr jnet large enongh to be easily distinguished free the noise (19) .
The sensitivity, ar the reoiprocal of threshold, xae
expressed as the Kodak neutral density filter number (N,D,No,) planed in the atimnlua 11ght path ; the higher the number, the loxer the threehold~ and the more sensitive the retina . 6-HD (dose expressed in aaeole of the hydrobromide per eye, Regis Chemioal Co .) xas dissolved in saline immediately before injection.
All experiments xere made on mature light-adapted albino rats
(280-350 gm) xhioh were housed normslly in the animal quarters and illuminated further in a xhite plsetio pan far 30 min under the maximum laboratory light (about 200 ft . oand]a) . After the intravitreaua injection of 10 pl saline or the drug solution into ether-aneethetised rate with a 30 Oa, needle and Hamilton syringe, the animals xere dark adapted immediately.
For the study
of the effeot of 6-HD (1 3uoole/eye) on the temporal oouree of dark adaptation, the rata xere anesthetiaed xith Nembutal (55 ~/kg~ i.p .), prepared under dim red light for ERO recording, and thresholds determined at intervals of time during dark adaptatia~ .
Otherxiae, the anieiale xsre dark adapted for 4-6 hours
before t!aeeholde xsre dster~ined .
Sinne preliminary results indicated that
the effeot of 6-iß on retinal sensitivity xae too drastic to esoluds the io-
Vol. 13, No . 1
17
6-Iiydroxydopamine and Dark Adaptation
volvement of photophemical mechanisms, the effeot of 6-iß on the rhodapain content xae also examined . abaarption at
5ßß
Rhodopeia oontent was measured by the decrease is
nm upon total blsaohing of a digitonin extract of retinal
tissue (19) and exvreeaed as percent of the dark-adapted saline-injected control value . Results and Disoueeion Aa aho~m in FIß . 1, the recovery of sensitivity of the saline injected
100 90 80
ae
~o
z .,
80 50 40 90
a
O A O
x a
ao lo
0
FIß . 1 Effect of 6-im (1 pmole/eye, solid) oo the temporal oouree of dark adaptation in the rat retina, Sensitivity (airole~ N-4-6) xaa eapreased as the äodak neutral density filter number (N .D.No.) planed in the atiaulua light path . Rhodopein pontent (aquare~ N~3-5) of saline control (open) a®rued as 100y6. Mean + S.E . control during the slop phase of dark adaptation is dependent on the regeneration of rhodopain (19) .
ßne pools of 6~ prevented both the regener-
ation of rhodopain and the recovery of sensitivity during the temparel noose of dark adaptation .
Following the injection to light,sdapted rate, xhioh
xere subsequently dark adapted for 4-6 hours, it xaa observed that one pools
18 of
8-Hydroxydopamine and Dark Adaptation Ô-HD
Vol. 13, No . 1
rendered the retina a]aoat nmeeponsive to the strongest light atiwli
and rednoed the rhodopain content to lA .s~ (TABIS 1) . Moreover, this effeot xaa highly done-dependent (FIG . 2) .
This drastic effect xaa prevented by
1 vole of Ka2S205, an antiaa3dant.
Iia2S205 itself had no effect on rhodopsin
oontent and only deoreased the sensitivity slightly (TAHIE 1) .
Ascorbio said
at 0.~ (0 .057 anoole) solution xhioh has generally been used to retard the oxidation of
6-HD,
had no beneficial effeot, xhile asoorbio aoid at 1 sole
partially prevented the 6-I~-induced effect . Aacorbio acid (1 ßoù) itself had no effeot on rhodopain content and decreased the sensitivity only slightly. 6-iß is readily oxidised in neutral aqueous aolutioo ae judged by the aLsost immediate discoloration .
Since Na 2S2O5 and aeaorbio acid (1 ample/10 pl)
prevented the discoloration of 6-I~ (1 nmole/10 pl) for apprasioately 10 and 4 days respectively, the difference in the effectiveness of the txo antio~ddante in preventing the 6-im-induced effect could be accounted for by their effeotiveneae in preventing the oxidation of 6-~. The molecular aechanis~o of action of 6-F~ in inducing the degeaeratioo of adrenergic nerve terminal ie still nnaertain.
6-Fm ie readily auto-aoddi :ed
to ample p-quinone (20-22) xhiah undergoes further transfarawtion to indo]iae and iodole derivatives (20~21~23) .
It has been proposed that the simple p-
quinone and indoline-indole derivatives undergo covalent binding xith pucLeophilic groups of biological macraaoleoules leading to a i~nctional impairment of the structure imrolved (21-2j) .
It hoe been long apeculsted that a con-
formational change in the photoreceptor protein (opein) resulting froei light stimuletioa ezposee nex active groups (24-26) .
It ie thought that the re-
generation of rhodopsin involves the covalent bidding of 11-oia retinal to the active groups of the opein (25~2T) .
The auddation products of Ô-HD could
undergo covalent binding xith these exposed active groups of opsin in the light-adapted retina, and thus prevent the regeneration of rhodopein and conooaitantly the recovery of retinal sensitivity.
Na 2,S 205 sad ascorbic acid
prevented the cod.dation of 6-F~, thus preventing its drastic effeot .
Although
Vol. 13, No. 1
8-Hydro~dopa,mine and Dark Adaptation
19
TARIE 1 Effeota of 6-® on retinal aensiti~i.tiea and rhodopein contents in the retim folloring administration to light~edapted rata and subsequently dark adapted for 4-6 hours
compound ga ~~M
(~o%ye)
(N.D,No,t+g,$.) E
-
(~,~
6-Hydros~dopa~iae
1
o,ti3 + 0,15
6-H>dYmgrdopamine in O.1K asocz~bio aoid aalina
1
0,59 + 0.23
6-H~drmcydopa~ins + aseorbio aoid
1 Eaoh
3.52 + 0,28
Asoorbio Aaid
1
6~ydrm~ydopamine + Na 25205
1 Eaoh
4 .59 + 0,08
xa 2s2o5
i
+ 0,04
~Oroôti(f)
H
10 100,0~ 5.9 8 19 14 5
10.5± 2.9 4 50.7+ 7 .5 5
6
100,Ot 4.6 5
3 .45 _ 0.32
5
85.4± 8,2 5
3 .24 + 0,16
6
100,3± il,o 4
ao effect of 5-,® on dopaminergio neurone in the retina is antioipated~thia effect might be masked by its drastic, non-epeoitio effect an rhodopein. The results of this etndy tend to support the hypothesis that the tao~ioity o! 6-im is mediated through the binding of its auddation prodnot(e) xith nucleophilio granpa of biological oaoroawleoul+as .
Vol. 13, No . 1
8-Hydroacydopamine and Dark Adaptation
20
z ci z
z m
F
7q
a 0 a0 a
F
m z
wm
0
0.188
0.181
0.588
0.511
0.716
L0
LOG D088 OF 6-HD (pmole)
FIG. 2 Effeota oß different doses of 6-HD (solid) on retinal aenaitivitea (oirole, N~6-19) and rhodopein cootenta (square, N-S-e) in the ratios iolla~iog administration of light-sdapted rats and anbaegnently dark adapted ßar 4-6 hours . AoknarledgmentThis work was partially anpported by Postdootaral Training Grant (MH 10695) to S.F.P, and by S11 Lilly & Co ., Reaearoh to Prevent Blindness, Inc . to L.T .G . The authors thank Hr . L. Tomey for technical aeeiataaoe, Referencea 1.
T . MALMFORS, Aota Phyaiol. Scand. ~ 99-100 (1963) .
2.
J. HAG(~NDAL and T. MALMFORS, Acta Phyaiol. Scand. ~, 58~56 (1965) .
3. A .M . LATIES and D. JACOHOWITZ, Anal . Reo. ~ 383-396 (1966) .
5
42-52 (1966) .
4.
B . EHIt~3ER, Irrveat. Ophthal.
5.
B. EHINGER, Z. Zellßoa~ach. 9~, 285-297 (1969) .
6.
A . AMES, III and D.A . POLLEN, J.Neura~h~eiol . ~ 424-442 (1969) .
7.
M. STRASCHILL and J. PSRWSIN, Pßlutaera Aroh, gurop, J. Phyaiol. 312,
45-54 (1969) .
Vol. 13, No . 1
8-Hydro~dopamine and Dark Adaptation
21
8.
K. KOJIHA,Y. MAJIMA, N. Z0 .SHIDA, U. OKOCAI, Y. NIIMI asd H. SûJIIâI, 24-27 (1965) . Jap. J. Ophthal.
9.
A . MDSTAKALLION, Acta Ophth. Snppl. 93 1-62 (1967) .
10,
69
5 .0 . KRAi+fER, Iereet . Ophthal.
1
438~52 (1971),
k
11, B .D . DRUJAN, J.H . DIAZ BOBGES and N . ALVERSZ, Life Soi, 473,477 (1965) . -
6
12 .
C.W . NICHOLS, D. JACOBWITZ and M. R0ITENSTEIN, Isveat . Ophthal. 642-646 (1967) . -
13 .
S .E .T . MALMOORS and H. THOENEN (eda .), 6-Fi >tine Neurosa , American S Lsevi.er, New Yor
14 .
C .O . GUTIBRREZ asd R.D . SPIGUSL, Viaion Rea . Suppl. ~, 161-170 (1971) .
15 .
J. VON HREDOid, S. BAY and N. ADAMS, Eap. Neurol. ~ 45-52 (1971) .
16 .
L.T . GRAHAM, JR, and S.F. POND, Aba . 5th Ist_ . Cones. Pharmacol., p. 97 (1972) .
17 . R.A . CONS, J. Gen . Phyaiol. 18,
1
m
anriss and Cateohola-
1267-1286 (1963) .
L.T . CRARAM, JR, asd S.F. PONG, ~. Neurol. 3~,6, 399-403 (1972) .
19 .
J.E . DOWLING, J. Ges. Phyaiol.
20.
S. SSNOH and B. WITKOP, J. Amer . Chem . Soc . 91, 6231235 (1959) .
~,
1287-1301 (1963) .
21 . A . SANER and H. THOSCtEN, Mol. Pharmaool. ~, 147-154 (1971) . 22 . R,N . AMMS, R. NhCREERY, L. HLANK and M. KAROLCZAK, Sur. J. Phermaool. 287-292 (1972) . 23 .
1
C.L . BLANK, P.T . KISSINGER and R .N . ADAMS, Sur. J. Pharmacol. 391-394 (1972) .
24 .
G . WALD and P .K . ~t.OWN, J, Gen . Phyeiol. 35, 797-822 (1952) .
25 .
J. HELLEQ, Bioohemiatry ~ 2914-2920 (1968),
26 .
C .H . RADDING and G . WALD, J. Gen . Phyaiol. 39, 909-922 (1956) .
27 .
H. AKHPAR, P .T . BIASES and P.H . DBtiüi[1RST, Chem. Comet.
1
s, 631-632 (1967) .
Lüe Sciences Vol . 13, pp . 15-21, 1973 . Printed in Great Britain
EFFECT OF 6-HYIROâ7mOPAMINB ON DARR ADAPTATION IN Tf~ RAT RETINA Schrs F. Pong and Lexie T. Graham, Jr . 9eotion of Neurobiology, The Institute of Peychiatrio Reaearoh~ Indiana University Medioal Center, Indianapolis, Indiana 46202 (Received 19 February 1973 ; in final form 9 Aprü 1973) SUl4~ARY 6-liydrozydopamine administered intravitreouaiy to Lightadapted rate appears to prevent the reocvery of retinal sensitivity during subsequent dark adaptation through its inter action xith the photochemical meohanisma . Adreaergio neurone, mainly containing dopamine, have been localised at the junction of the retinal inner nuclear and inner pleziform layers of all species so far investigated by the fluo~reaoenoe technique (1-5) .
Neuro
phyaiological studies have shown that ezogenously applied dopamine can inhibit the firing of retinal ganglion cells (6,7) . The ensymes responsible for the eetabolio degradation of aateoholaminea (8,9) and the uptabe and release of dopamine from the cat ratios follaring appropriate light stimulation have been demonstrated (10) . Thus, the inhibitory neurotransmitter role of retinal dopamine is strongly implicated .
However, littù direct information con-
cerning the functional role of dopamine in the retina ie available .
Attempts
to correlate the dopamine content, Wonoamine o~xidaae and aateohol-0 ..eethyl traneferaae activities to the state of adaptation have been reported, but not xithout some conflicting results (8,11,12) .
It is of interest to ants that
anitwls treated xith raserpine, a cateoholamioe-depleting agent, ezEsibit photophobia xhioh eight bs due to a blockade of aminergio inhibitory meohaniaes (1) . It has been xell established that 6-hydroxydopamine (6-HD) causes a selective and long-lasting daganeration of adrsnergio nerve tersinala and has been xidely employed recently as a phara~acologioal tool to study the fractions 15
18
6-Hydroxydopamine and Dark Adaptation
of adreoergio neurons (13) .
Vol . 13, No . 1
Due to its remarkable ohemical eympatheotaoy
property and to the feasibility of detecting changes provoked by pharmacological manipulation of retinal neurone by monitoring the eleotroretinogram (ERG) (1lt-16), and because of our interest in functional correlatione of neurotransmitters in retina, the effect of 6-HD on the ERG threshold of the rat retina was studied, Ptethode The appearanoe of the brave of the ERG xae used as a measure of retinal sensitivity and xae determined under experimental conditions similar to that used by Cone (17) as described previously (18),
The threshold ie the mini~nuo
Light stimulus neoeeeary to evoke a just-peroeptib]s response in the BR,G . Depending on the noise of the partioular preparation, the response aeleoted xae a potential of 10 to 20 pvr jnet large enongh to be easily distinguished free the noise (19) .
The sensitivity, ar the reoiprocal of threshold, xae
expressed as the Kodak neutral density filter number (N,D,No,) planed in the atimnlua 11ght path ; the higher the number, the loxer the threehold~ and the more sensitive the retina . 6-HD (dose expressed in aaeole of the hydrobromide per eye, Regis Chemioal Co .) xas dissolved in saline immediately before injection.
All experiments xere made on mature light-adapted albino rats
(280-350 gm) xhioh were housed normslly in the animal quarters and illuminated further in a xhite plsetio pan far 30 min under the maximum laboratory light (about 200 ft . oand]a) . After the intravitreaua injection of 10 pl saline or the drug solution into ether-aneethetised rate with a 30 Oa, needle and Hamilton syringe, the animals xere dark adapted immediately.
For the study
of the effeot of 6-HD (1 3uoole/eye) on the temporal oouree of dark adaptation, the rata xere anesthetiaed xith Nembutal (55 ~/kg~ i.p .), prepared under dim red light for ERO recording, and thresholds determined at intervals of time during dark adaptatia~ .
Otherxiae, the anieiale xsre dark adapted for 4-6 hours
before t!aeeholde xsre dster~ined .
Sinne preliminary results indicated that
the effeot of 6-iß on retinal sensitivity xae too drastic to esoluds the io-
Vol. 13, No . 1
17
6-Iiydroxydopamine and Dark Adaptation
volvement of photophemical mechanisms, the effeot of 6-iß on the rhodapain content xae also examined . abaarption at
5ßß
Rhodopeia oontent was measured by the decrease is
nm upon total blsaohing of a digitonin extract of retinal
tissue (19) and exvreeaed as percent of the dark-adapted saline-injected control value . Results and Disoueeion Aa aho~m in FIß . 1, the recovery of sensitivity of the saline injected
100 90 80
ae
~o
z .,
80 50 40 90
a
O A O
x a
ao lo
0
FIß . 1 Effect of 6-im (1 pmole/eye, solid) oo the temporal oouree of dark adaptation in the rat retina, Sensitivity (airole~ N-4-6) xaa eapreased as the äodak neutral density filter number (N .D.No.) planed in the atiaulua light path . Rhodopein pontent (aquare~ N~3-5) of saline control (open) a®rued as 100y6. Mean + S.E . control during the slop phase of dark adaptation is dependent on the regeneration of rhodopain (19) .
ßne pools of 6~ prevented both the regener-
ation of rhodopain and the recovery of sensitivity during the temparel noose of dark adaptation .
Following the injection to light,sdapted rate, xhioh
xere subsequently dark adapted for 4-6 hours, it xaa observed that one pools
18 of
8-Hydroxydopamine and Dark Adaptation Ô-HD
Vol. 13, No . 1
rendered the retina a]aoat nmeeponsive to the strongest light atiwli
and rednoed the rhodopain content to lA .s~ (TABIS 1) . Moreover, this effeot xaa highly done-dependent (FIG . 2) .
This drastic effect xaa prevented by
1 vole of Ka2S205, an antiaa3dant.
Iia2S205 itself had no effect on rhodopsin
oontent and only deoreased the sensitivity slightly (TAHIE 1) .
Ascorbio said
at 0.~ (0 .057 anoole) solution xhioh has generally been used to retard the oxidation of
6-HD,
had no beneficial effeot, xhile asoorbio aoid at 1 sole
partially prevented the 6-I~-induced effect . Aacorbio acid (1 ßoù) itself had no effeot on rhodopain content and decreased the sensitivity only slightly. 6-iß is readily oxidised in neutral aqueous aolutioo ae judged by the aLsost immediate discoloration .
Since Na 2S2O5 and aeaorbio acid (1 ample/10 pl)
prevented the discoloration of 6-I~ (1 nmole/10 pl) for apprasioately 10 and 4 days respectively, the difference in the effectiveness of the txo antio~ddante in preventing the 6-im-induced effect could be accounted for by their effeotiveneae in preventing the oxidation of 6-~. The molecular aechanis~o of action of 6-F~ in inducing the degeaeratioo of adrenergic nerve terminal ie still nnaertain.
6-Fm ie readily auto-aoddi :ed
to ample p-quinone (20-22) xhiah undergoes further transfarawtion to indo]iae and iodole derivatives (20~21~23) .
It has been proposed that the simple p-
quinone and indoline-indole derivatives undergo covalent binding xith pucLeophilic groups of biological macraaoleoules leading to a i~nctional impairment of the structure imrolved (21-2j) .
It hoe been long apeculsted that a con-
formational change in the photoreceptor protein (opein) resulting froei light stimuletioa ezposee nex active groups (24-26) .
It ie thought that the re-
generation of rhodopsin involves the covalent bidding of 11-oia retinal to the active groups of the opein (25~2T) .
The auddation products of Ô-HD could
undergo covalent binding xith these exposed active groups of opsin in the light-adapted retina, and thus prevent the regeneration of rhodopein and conooaitantly the recovery of retinal sensitivity.
Na 2,S 205 sad ascorbic acid
prevented the cod.dation of 6-F~, thus preventing its drastic effeot .
Although
Vol. 13, No. 1
8-Hydro~dopa,mine and Dark Adaptation
19
TARIE 1 Effeota of 6-® on retinal aensiti~i.tiea and rhodopein contents in the retim folloring administration to light~edapted rata and subsequently dark adapted for 4-6 hours
compound ga ~~M
(~o%ye)
(N.D,No,t+g,$.) E
-
(~,~
6-Hydros~dopa~iae
1
o,ti3 + 0,15
6-H>dYmgrdopamine in O.1K asocz~bio aoid aalina
1
0,59 + 0.23
6-H~drmcydopa~ins + aseorbio aoid
1 Eaoh
3.52 + 0,28
Asoorbio Aaid
1
6~ydrm~ydopamine + Na 25205
1 Eaoh
4 .59 + 0,08
xa 2s2o5
i
+ 0,04
~Oroôti(f)
H
10 100,0~ 5.9 8 19 14 5
10.5± 2.9 4 50.7+ 7 .5 5
6
100,Ot 4.6 5
3 .45 _ 0.32
5
85.4± 8,2 5
3 .24 + 0,16
6
100,3± il,o 4
ao effect of 5-,® on dopaminergio neurone in the retina is antioipated~thia effect might be masked by its drastic, non-epeoitio effect an rhodopein. The results of this etndy tend to support the hypothesis that the tao~ioity o! 6-im is mediated through the binding of its auddation prodnot(e) xith nucleophilio granpa of biological oaoroawleoul+as .
Vol. 13, No . 1
8-Hydroacydopamine and Dark Adaptation
20
z ci z
z m
F
7q
a 0 a0 a
F
m z
wm
0
0.188
0.181
0.588
0.511
0.716
L0
LOG D088 OF 6-HD (pmole)
FIG. 2 Effeota oß different doses of 6-HD (solid) on retinal aenaitivitea (oirole, N~6-19) and rhodopein cootenta (square, N-S-e) in the ratios iolla~iog administration of light-sdapted rats and anbaegnently dark adapted ßar 4-6 hours . AoknarledgmentThis work was partially anpported by Postdootaral Training Grant (MH 10695) to S.F.P, and by S11 Lilly & Co ., Reaearoh to Prevent Blindness, Inc . to L.T .G . The authors thank Hr . L. Tomey for technical aeeiataaoe, Referencea 1.
T . MALMFORS, Aota Phyaiol. Scand. ~ 99-100 (1963) .
2.
J. HAG(~NDAL and T. MALMFORS, Acta Phyaiol. Scand. ~, 58~56 (1965) .
3. A .M . LATIES and D. JACOHOWITZ, Anal . Reo. ~ 383-396 (1966) .
5
42-52 (1966) .
4.
B . EHIt~3ER, Irrveat. Ophthal.
5.
B. EHINGER, Z. Zellßoa~ach. 9~, 285-297 (1969) .
6.
A . AMES, III and D.A . POLLEN, J.Neura~h~eiol . ~ 424-442 (1969) .
7.
M. STRASCHILL and J. PSRWSIN, Pßlutaera Aroh, gurop, J. Phyaiol. 312,
45-54 (1969) .
Vol. 13, No . 1
8-Hydro~dopamine and Dark Adaptation
21
8.
K. KOJIHA,Y. MAJIMA, N. Z0 .SHIDA, U. OKOCAI, Y. NIIMI asd H. SûJIIâI, 24-27 (1965) . Jap. J. Ophthal.
9.
A . MDSTAKALLION, Acta Ophth. Snppl. 93 1-62 (1967) .
10,
69
5 .0 . KRAi+fER, Iereet . Ophthal.
1
438~52 (1971),
k
11, B .D . DRUJAN, J.H . DIAZ BOBGES and N . ALVERSZ, Life Soi, 473,477 (1965) . -
6
12 .
C.W . NICHOLS, D. JACOBWITZ and M. R0ITENSTEIN, Isveat . Ophthal. 642-646 (1967) . -
13 .
S .E .T . MALMOORS and H. THOENEN (eda .), 6-Fi >tine Neurosa , American S Lsevi.er, New Yor
14 .
C .O . GUTIBRREZ asd R.D . SPIGUSL, Viaion Rea . Suppl. ~, 161-170 (1971) .
15 .
J. VON HREDOid, S. BAY and N. ADAMS, Eap. Neurol. ~ 45-52 (1971) .
16 .
L.T . GRAHAM, JR, and S.F. POND, Aba . 5th Ist_ . Cones. Pharmacol., p. 97 (1972) .
17 . R.A . CONS, J. Gen . Phyaiol. 18,
1
m
anriss and Cateohola-
1267-1286 (1963) .
L.T . CRARAM, JR, asd S.F. PONG, ~. Neurol. 3~,6, 399-403 (1972) .
19 .
J.E . DOWLING, J. Ges. Phyaiol.
20.
S. SSNOH and B. WITKOP, J. Amer . Chem . Soc . 91, 6231235 (1959) .
~,
1287-1301 (1963) .
21 . A . SANER and H. THOSCtEN, Mol. Pharmaool. ~, 147-154 (1971) . 22 . R,N . AMMS, R. NhCREERY, L. HLANK and M. KAROLCZAK, Sur. J. Phermaool. 287-292 (1972) . 23 .
1
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