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Labeled noradrenaline release from rat cerebral cortex following electrical stimulation of locus coeruleus
384
Btam Resealth, 106 (1976) 384-389 .(~, Elsewer Sclentrfic Pubhshmg Company, Amsterdam Printed m The Netherland~
Labeled noradrenaline release from rat cerebral cortex following electrical stimulation of locus coeruleus
CHIKAKO TANAKA, CHIYOKO INAGAKI AND HIROSHI FUJIWARA Department of Pharmacology, Faculty of Medicine, Kyoto Universtty, Sankyo-ku, Kyoto 606 (Japan)
(Accepted January 20th, 1976)
Histochemical studies reveal that the locus coeruleus (LC) in the rat consists of cell bodies of noradrenaline (NA) containing neurons which project to the cerebral cortex 1,1°,12,17. Recently, an increase in N A metabolites as well as turnover and a decrease of N A content in the forebrain were found to occur as a result of electrical stimulation of the LC in this same species s,9,1a. These findings, however, provide only indirect evidence for NA release from the cerebral cortex with LC stimulation. For collecting transmitters or presumed transmitters in vivo, various techniques such as the push-pull cannula 5, ventricular perfusion 14 and the cortical cup technique T M have been used. With the latter technique, released substances from the limited cortex can be collected 13. Brenells 4 has demonstrated the release of labeled N A from the rabbit olfactory bulbs upon stimulation of the medial olfactory tract. Studies on the released NA collected from the cerebral cortex following stimulation of a major source such as LC have never been documented and such an approach was attempted by our group in vivo. Thirty-one Wistar male rats, weighing 200-240 g, were anesthetized with either urethane (1.6 g/kg) or pentobarbital sodium (30 mg/kg) and the released labeled N A was collected using the cortical cup technique. Rats were fixed in a stereotaxic apparatus and the skull was removed, usually on both sides of the midline, from 2 m m anterior to iambda to 2 m m anterior to bregma. The overlying dura was reflected and the cerebral cortex was exposed. Two cylindrical plastic cups (4.5 m m in internal diameter, 8 m m m height) were placed on the exposed cortex and sealed with surgical adhesive. Each cup was filled with modified Krebs phosphate solution and contained E D T A (disodium salt 10 mg/l) and ascorbic acid (20 mg/l). This solution was gassed with 95 9/00 02-5 9/ooCO2 at 37 °C before addition to the cups. The cerebral cortex was incubated for 1 h with [1,7-3H]noradrenaline ([3H]NA, 8.2 Ci/mole) dissolved in the modified Krebs phosphate solution. After incubation, the medium was removed and the cortex in the cup was washed five times with a fresh solution. The cups were then filled with 0.05 ml of the solution containing 10 -4 M desipramine to reduce the reuptake of NA. Samples were collected at 10 min intervals. Total radioactivity originating from [aH]NA was counted in a Packard Tri-Carb liquid scintillation
385 spectrometer for a 10 min period and this radioactivity was expressed as the number of disintegrations per minute (d.p.m.) For stimulation of LC, bipolar electrodes 0.3 mm in diameter and made from stainless steel wires were inserted into the LC. The stereotaxic coordinates were determined in the initial experiment by the fluorescence histochemical technique of Falck and Hillarp 6 and were confirmed after every experiment by routine histology. The stimuli composed of monophasic pulses of 2 msec and 4 V were applied to the LC. Frequency and stimulation time varied between 5-30 Hz and 2-8 min. During the experiments, body temperature was maintained at 36.5-37.5 °C. Spontaneous efflux of total radioactivity declined in the multiphasic time course, rapidly at first and slowly later on. In the first 50 min, the decline rates of spontaneous efflux were k 4- S.E. = 0.074 4- 0.003 min -1 in the rat anesthetized with urethane (urethane-rats) and k ± S.E. = 0.066 4- 0.003 min -1 in the animals anesthetized with pentobarbital sodium (pentobarbital-rats). The efflux of total radioactivity was significantly greater in the urethane-rats (Table I). Total radioactivities of released [SH]NA and its metabolites at the 180 min period were 1344 4106 d.p.m, in the urethane-rats and 748 4- 60 d.p.m, in the pentobarbital-rats respectively. These data indicate that the spontaneous efflux of total activity was significantly suppressed in the pentobarbital-rats as compared with that in the urethane-rats. The LC stimulated release of [aH]NA and its metabolites from the cerebral cortex was unmeasurable in the absence of desipramine; therefore, desipramine 10-4 M was added to the solution in the cups in all experiments. The LC stimulated release was dependent on the frequency and duration of stimulation and stimulation at a frequency of 20 Hz for periods of 2 min resulted in a significant increase in release of [aH]NA and its metabolites (Fig. 1). Even with a frequency of 5 Hz, the release was measurable and linearly increased with 10-20 Hz stimulation (Fig. 2). There was an even greater increase with a 30 Hz stimulation. No linear relationship between stimulation time and release of [aH]NA and its metabolites was seen with stimulation at 2, 5 and 8 min. Five tracking experiments were done. The effective sites for ipsilateral release were confined within very narrow limits, the stereotaxic coordinates posterior 1.0-1.5 mm, lateral 1.0-1.2 mm and vertical 6.7-7.3 mm from the surface of the skull. HistoTABLE I DECLINE RATE AND TOTAL RADIOACTIVITYOF SPONTANEOUSRELEASE OF [aH]NA A~D rrs METABOLITE$ FROM TIlE CEREBRAL CORTEX Each value is the mean of 14 experiments on 7 animals.
Urethane-rats Pentobarbital-rats
rapid decline k i S.E. min -1
slow decline k qS.E. mill -1
d.p.m, at 180 min period
0.074 4- 0.003 0.066 4- 0.003
0.0079 -4- 0.0005 0.0049 4- 0.0003*
1344 -4- 106 748 q- 60*
* Significantly different f r o m value in the urethane-rats at P < 0.001 (Student's t-test).
386 5
'o
20
10
30Hz
20
2
x
E
±~
!
10 I
L3 L- 1 I"1i...
2
i
I--.. I I.. I i
=--r--L..r_ I
|
I
0
60
I
|
120 180 Time (rain)
I
I
240
300
Fig. 1. Release of [aH]noradrenatine and its metabohtes from the ipsitateral cortex following urnlateral electrical stimulation of the locus coeruleus (lateral 1.2 mm, posterior 1+2 mm, vertical 6 8 ram) in rats anesthetized with pentobarbital sodium 30 mg/kg. The solid line represents total radioactivities released from the ipsilateral cortex and the dotted line those from the contralateral cortex. Arrows indicate electrical stimulation (4 V, 20 Hz, 2 msec, 2 min).
logical examination of the stimulation sites demonstrated that these corresponded to the LC and the proximal area of dorsal bundle. A typical case of how tracking was done along the dorsoventral axis at the electrode position in posterior 1.5 m m and lateral 1.2 mm, is illustrated in Fig. 3. [aH]NA and its metabolites were released from the ipsilateral cortex at the location of vertical 6.8 mm, from the bilateral cortex at vertical 7.3 m m and from the contralateral cortex at vertical 7.8 mm. The most 2-
J
t~
t~1
....30 Hz Purses per second
Fig. 2. Relationship between the frequencies of stimulation and the ratio of LC-stimutated release
(Rst) to spontaneous release (Rsp) of [aH]noradrenaline and its metabolites from the ipsilateral cortex. Each point represents the mean of 4 experiments.
387
2G lC O
E cL 3
'1o
"~20 Z
lO
--1.° I
'--,_..~_._.~--,--~ ...........
~--, I
~'~
i'--7----'i
I
I
I
l
120
180
2/,0
300
Time (ram) Fig. 3. Release of [gH]noradrenaline and its metabolites from the cerebral cortex following unilateral electrical stimulation of various sites of vertical 5.8, 6.8, 7.3, 7.8, and 8.8 mm along the dorsoventral axis at the electrode position in posterior 1.5 and lateral 1.2. The solid line represents total radioactivities released from the ipsilateral cortex and the dotted line those from the contralateral cortex. Arrows indicate electrical stimulation (4 V, 20 Hz, 2 msec, 2 min).
effective sites for ipsilateral release were found in the anterior pole of LC and those from contralateral release in the ventral part of LC and the region rostral and medial to LC. The present study has demonstrated that spontaneous efflux of total radioactivity declines in a multiphasic time course. The decline rates of release from the rat cerebral cortex were almost similar to those from the olfactory bulbs in the rabbit under urethane anesthesia4. The application of pentobarbital anesthesia rather than urethane significantly suppressed spontaneous etttux in our experiment. As the output of acetylcholine from the surface of cortex reportedly depends on the depth of anesthesia and is approximately proportional to the spontaneous cortical activityza, further experiments are required to clarify whether differences in the spontaneous release of cortical NA between the urethane-rats and the pentobarbital-rats depends on the depth of anesthesia or the mode of action of these drugs. Since the radioactivity attributed to unchanged NA was reported to be initially 91 ~o and 6 0 ~ after 4.5 h of sample collection in the rabbit olfactory bulbs 4, total radioactivities of sample collections from the rat cerebral cortex are considered to be a good reflection of the radioactivity of unchanged NA. LC stimulation-induced release of [aH]NA and its metabolites was demonstrated in this study to be frequency-related and to increase following the blockade of reuptake of NA. Of great interest are the findings that the cortical NA release was
388 most d e p e n d e n t on the locahzat~on o f stimulation electrodes and that stimulation ol the ventral part o f LC and the region rostral a n d medial to LC reduced release o f [3H]NA and its m e t a b o h t e s n o t only from the stimulated side but also from the nons t i m u l a t e d s~de. It has been stated that the cortical p r o j e c t i o n o f LC ~s ~psdateral, by reason o f N A depletion m the Jpsilateral cortex following umlateral destruction o f LC2,3,7,17. K o r f et al. 'q a n d W a l t e r a n d Eccleston t8 s u p p o r t e d the t h e o r y o f ipsdateral cortical mnervation. They considered slight changes o f N A and ~ts m e t a b o h t e s m the c o n t r a l a t e r a l cortex as p r o b a b l y due to m & r e c t s t i m u l a t i o n as a result o f spread o f c u r r e n t from the electrode. Shinuzu et al. xe', however, d e m o n s t r a t e d w~th their m o r p h o l o g i c a l ewdence that the u n d a t e r a l lesion to L C produces a considerable n u m b e r o f d e g e n e r a t e d fibers n o t only in the ipsdateral cortex b u t also m the c o n t r a lateral cortex. Rets and Ross a5 have also f o u n d that d o p a m m e - f l - h y d r o x y l a s e act w i t y in the rat r e m a i n e d lowered in the ipsilateral cortex for up to 45 days, and also fell transiently for 7 days m the c o n t r a l a t e r a l cortex following LC destruction. The present w o r k revealed t h a t [3H]NA release from the c o n t r a l a t e r a l cortex occurred following stimulation o f the ventral p a r t o f LC a n d the region rostral a n d medml to LC. O u r findings that s t l m u l a h o n o f the LC results m N A release not only from the lpsdateral cortex but also f r o m the c o n t r a l a t e r a l cortex, are considered to s u p p o r t the hypothesis o f a bilateral cortical innervation from the LC. F u r t h e r experiments, e.g. cutting o f the c o n t r a l a t e r a l d o r s a l N A bundle, are necessary to rule out direct or indirect increase m ~mpulse flow t h r o u g h the c o n t r a l a t e r a l LC. This w o r k was s u p p o r t e d by g r a n t N o . 944041 f r o m the Ministry o f E d u c a h o n , Japan. The a u t h o r s express their gratitude to Prof. S. T a k a o r i for p e r t i n e n t advice a n d to M. O h a r a for assistance with the manuscript.
1 ANDI~N,N.-E., DAHLSTROM,A., FUXE, K , OLSON, L., AND UNGERSTEDT,U., Ascending monoamine neurons to the telencephalon and &encephalon, .4cta physiol, scand., 67 (1966) 313-326 2 ANLEZARK,G., CROW, J., AND GREENWAY,A., Impaired learning and decreased corhcal norepinephrine after bdateral locus coeruleus lesions, Scwnce, 181 (1973) 682-684. 3 ARBUTHNOTT,G W , CROW, T. J., FuxE, K., OLSON, L., AND UNGERSTEDT,U., Depletion of catecholamines in vivo induced by electrical stimulation of central monoamine pathways, Brain Research, 24 (1970) 471-483. 4 BRENELLS,A. B., Spontaneous and neurally evoked release of labelled noradrenahne from rabbit olfactory bulbs in vivo, J. Physiol. (Lond.), 240 (1974) 279-293 5 GADDUM,J. H., Push-pull cannulae, J Physiol. (Lond.), 155 (1961) 1-2. 6 FALCK,B., HILLARP,N -A., THIEME,G., ANDTOR, A., Fluorescence of catecholammes and related compounds condensed wxth formaldehyde, J. Histochem. Cytochem., 10 (1962) 348-354. 7 KOBAYASHI,R. M., PALKOVlTS,M., KOPIN, I. J., AND JACOBOWlTZ,D. M., B~ochem~cal mapping of noradrenergic nerves arising from the rat locus coeruleus, Brain Research, 77 (1974) 264-269. 8 KORF,J , AGHAJANIAN,G. K , AND ROTH, R. H., Stimulation and destruction of the locus coeruleus opposite effects on 3-methoxy-4-hydroxyphenylglycol sulfate levels m the rat cerebral cortex, Europ. J. PharmacoL, 2l (1973) 305-310. 9 KORF,J., ROTH, R. H., AND AGHAJANIAN,G. K , Alterations m turnover and endogenous levels of norepinephrine m cerebral cortex following electrical stimulation and acute axotomy of cerebral noradrenergic pathways, Europ. J. Pharmacol., 23 (1973) 276-282.
389 10 LoIzou, L. A., Projectlon of the nucleus locus coeruleus in the albino rat, Brain Research, 15 (1969) 563-566. II MACINTOSH, F. C., AND OBORIN, P. E., Release of acetylcholine from intact cerebral cortex, Abstr. X1X Int. Physiol. Congr., (1953) 580. 12 MAEDA, T., ET SHIMIZU, N., Projections ascendantes du locus coeruleus et d'autres neurones ammergiques pontlques au nlveau du prosenc6phale du rat, Brain Research, 36 (1972) 19-35. 13 MITCHELL,J. E., The spontaneous and evoked release of acetylcholine from the cerebral cortex, J. Physiol. (Lond.), 165 (1963) 98-116. 14 PORTIG, P. J. AND VOGT, M., Search for substances released on stimulation of the caudate nucleus m the cat, J. Physiol. (Lond.), 186 0967) 131-132. 15 REIS, D., AND ROSS, R,, Dynamic changes in brain dopamine-fl-hydroxylase activity during anterograde and retrograde reactions to injury of central noradrenergic axons, Brain Research, 57 (1973) 307-326. 16 SHIMIZU, N., OHNISHI, S., TOHYAMA,M., AND MAEDA, T., Demonstration by degeneration silver method of the ascending projection from the locus co~ruleus, Exp. Brain Res., 21 (1974) 181-192. 17 UNGERSTEDT,U., Stereotaxic mapping of the monoamine pathways in the rat brain, .4cta physiol. scand., Suppl. 367 (1971) 1-48. 18 WALTER, D. S., AND ECCLESTON,D., Increase of noradrenallne metabohsm following electrical stimulation of the locus coeruleus in the rat, J. Neuroehem., 21 (1973) 281-289.
Btam Resealth, 106 (1976) 384-389 .(~, Elsewer Sclentrfic Pubhshmg Company, Amsterdam Printed m The Netherland~
Labeled noradrenaline release from rat cerebral cortex following electrical stimulation of locus coeruleus
CHIKAKO TANAKA, CHIYOKO INAGAKI AND HIROSHI FUJIWARA Department of Pharmacology, Faculty of Medicine, Kyoto Universtty, Sankyo-ku, Kyoto 606 (Japan)
(Accepted January 20th, 1976)
Histochemical studies reveal that the locus coeruleus (LC) in the rat consists of cell bodies of noradrenaline (NA) containing neurons which project to the cerebral cortex 1,1°,12,17. Recently, an increase in N A metabolites as well as turnover and a decrease of N A content in the forebrain were found to occur as a result of electrical stimulation of the LC in this same species s,9,1a. These findings, however, provide only indirect evidence for NA release from the cerebral cortex with LC stimulation. For collecting transmitters or presumed transmitters in vivo, various techniques such as the push-pull cannula 5, ventricular perfusion 14 and the cortical cup technique T M have been used. With the latter technique, released substances from the limited cortex can be collected 13. Brenells 4 has demonstrated the release of labeled N A from the rabbit olfactory bulbs upon stimulation of the medial olfactory tract. Studies on the released NA collected from the cerebral cortex following stimulation of a major source such as LC have never been documented and such an approach was attempted by our group in vivo. Thirty-one Wistar male rats, weighing 200-240 g, were anesthetized with either urethane (1.6 g/kg) or pentobarbital sodium (30 mg/kg) and the released labeled N A was collected using the cortical cup technique. Rats were fixed in a stereotaxic apparatus and the skull was removed, usually on both sides of the midline, from 2 m m anterior to iambda to 2 m m anterior to bregma. The overlying dura was reflected and the cerebral cortex was exposed. Two cylindrical plastic cups (4.5 m m in internal diameter, 8 m m m height) were placed on the exposed cortex and sealed with surgical adhesive. Each cup was filled with modified Krebs phosphate solution and contained E D T A (disodium salt 10 mg/l) and ascorbic acid (20 mg/l). This solution was gassed with 95 9/00 02-5 9/ooCO2 at 37 °C before addition to the cups. The cerebral cortex was incubated for 1 h with [1,7-3H]noradrenaline ([3H]NA, 8.2 Ci/mole) dissolved in the modified Krebs phosphate solution. After incubation, the medium was removed and the cortex in the cup was washed five times with a fresh solution. The cups were then filled with 0.05 ml of the solution containing 10 -4 M desipramine to reduce the reuptake of NA. Samples were collected at 10 min intervals. Total radioactivity originating from [aH]NA was counted in a Packard Tri-Carb liquid scintillation
385 spectrometer for a 10 min period and this radioactivity was expressed as the number of disintegrations per minute (d.p.m.) For stimulation of LC, bipolar electrodes 0.3 mm in diameter and made from stainless steel wires were inserted into the LC. The stereotaxic coordinates were determined in the initial experiment by the fluorescence histochemical technique of Falck and Hillarp 6 and were confirmed after every experiment by routine histology. The stimuli composed of monophasic pulses of 2 msec and 4 V were applied to the LC. Frequency and stimulation time varied between 5-30 Hz and 2-8 min. During the experiments, body temperature was maintained at 36.5-37.5 °C. Spontaneous efflux of total radioactivity declined in the multiphasic time course, rapidly at first and slowly later on. In the first 50 min, the decline rates of spontaneous efflux were k 4- S.E. = 0.074 4- 0.003 min -1 in the rat anesthetized with urethane (urethane-rats) and k ± S.E. = 0.066 4- 0.003 min -1 in the animals anesthetized with pentobarbital sodium (pentobarbital-rats). The efflux of total radioactivity was significantly greater in the urethane-rats (Table I). Total radioactivities of released [SH]NA and its metabolites at the 180 min period were 1344 4106 d.p.m, in the urethane-rats and 748 4- 60 d.p.m, in the pentobarbital-rats respectively. These data indicate that the spontaneous efflux of total activity was significantly suppressed in the pentobarbital-rats as compared with that in the urethane-rats. The LC stimulated release of [aH]NA and its metabolites from the cerebral cortex was unmeasurable in the absence of desipramine; therefore, desipramine 10-4 M was added to the solution in the cups in all experiments. The LC stimulated release was dependent on the frequency and duration of stimulation and stimulation at a frequency of 20 Hz for periods of 2 min resulted in a significant increase in release of [aH]NA and its metabolites (Fig. 1). Even with a frequency of 5 Hz, the release was measurable and linearly increased with 10-20 Hz stimulation (Fig. 2). There was an even greater increase with a 30 Hz stimulation. No linear relationship between stimulation time and release of [aH]NA and its metabolites was seen with stimulation at 2, 5 and 8 min. Five tracking experiments were done. The effective sites for ipsilateral release were confined within very narrow limits, the stereotaxic coordinates posterior 1.0-1.5 mm, lateral 1.0-1.2 mm and vertical 6.7-7.3 mm from the surface of the skull. HistoTABLE I DECLINE RATE AND TOTAL RADIOACTIVITYOF SPONTANEOUSRELEASE OF [aH]NA A~D rrs METABOLITE$ FROM TIlE CEREBRAL CORTEX Each value is the mean of 14 experiments on 7 animals.
Urethane-rats Pentobarbital-rats
rapid decline k i S.E. min -1
slow decline k qS.E. mill -1
d.p.m, at 180 min period
0.074 4- 0.003 0.066 4- 0.003
0.0079 -4- 0.0005 0.0049 4- 0.0003*
1344 -4- 106 748 q- 60*
* Significantly different f r o m value in the urethane-rats at P < 0.001 (Student's t-test).
386 5
'o
20
10
30Hz
20
2
x
E
±~
!
10 I
L3 L- 1 I"1i...
2
i
I--.. I I.. I i
=--r--L..r_ I
|
I
0
60
I
|
120 180 Time (rain)
I
I
240
300
Fig. 1. Release of [aH]noradrenatine and its metabohtes from the ipsitateral cortex following urnlateral electrical stimulation of the locus coeruleus (lateral 1.2 mm, posterior 1+2 mm, vertical 6 8 ram) in rats anesthetized with pentobarbital sodium 30 mg/kg. The solid line represents total radioactivities released from the ipsilateral cortex and the dotted line those from the contralateral cortex. Arrows indicate electrical stimulation (4 V, 20 Hz, 2 msec, 2 min).
logical examination of the stimulation sites demonstrated that these corresponded to the LC and the proximal area of dorsal bundle. A typical case of how tracking was done along the dorsoventral axis at the electrode position in posterior 1.5 m m and lateral 1.2 mm, is illustrated in Fig. 3. [aH]NA and its metabolites were released from the ipsilateral cortex at the location of vertical 6.8 mm, from the bilateral cortex at vertical 7.3 m m and from the contralateral cortex at vertical 7.8 mm. The most 2-
J
t~
t~1
....30 Hz Purses per second
Fig. 2. Relationship between the frequencies of stimulation and the ratio of LC-stimutated release
(Rst) to spontaneous release (Rsp) of [aH]noradrenaline and its metabolites from the ipsilateral cortex. Each point represents the mean of 4 experiments.
387
2G lC O
E cL 3
'1o
"~20 Z
lO
--1.° I
'--,_..~_._.~--,--~ ...........
~--, I
~'~
i'--7----'i
I
I
I
l
120
180
2/,0
300
Time (ram) Fig. 3. Release of [gH]noradrenaline and its metabolites from the cerebral cortex following unilateral electrical stimulation of various sites of vertical 5.8, 6.8, 7.3, 7.8, and 8.8 mm along the dorsoventral axis at the electrode position in posterior 1.5 and lateral 1.2. The solid line represents total radioactivities released from the ipsilateral cortex and the dotted line those from the contralateral cortex. Arrows indicate electrical stimulation (4 V, 20 Hz, 2 msec, 2 min).
effective sites for ipsilateral release were found in the anterior pole of LC and those from contralateral release in the ventral part of LC and the region rostral and medial to LC. The present study has demonstrated that spontaneous efflux of total radioactivity declines in a multiphasic time course. The decline rates of release from the rat cerebral cortex were almost similar to those from the olfactory bulbs in the rabbit under urethane anesthesia4. The application of pentobarbital anesthesia rather than urethane significantly suppressed spontaneous etttux in our experiment. As the output of acetylcholine from the surface of cortex reportedly depends on the depth of anesthesia and is approximately proportional to the spontaneous cortical activityza, further experiments are required to clarify whether differences in the spontaneous release of cortical NA between the urethane-rats and the pentobarbital-rats depends on the depth of anesthesia or the mode of action of these drugs. Since the radioactivity attributed to unchanged NA was reported to be initially 91 ~o and 6 0 ~ after 4.5 h of sample collection in the rabbit olfactory bulbs 4, total radioactivities of sample collections from the rat cerebral cortex are considered to be a good reflection of the radioactivity of unchanged NA. LC stimulation-induced release of [aH]NA and its metabolites was demonstrated in this study to be frequency-related and to increase following the blockade of reuptake of NA. Of great interest are the findings that the cortical NA release was
388 most d e p e n d e n t on the locahzat~on o f stimulation electrodes and that stimulation ol the ventral part o f LC and the region rostral a n d medial to LC reduced release o f [3H]NA and its m e t a b o h t e s n o t only from the stimulated side but also from the nons t i m u l a t e d s~de. It has been stated that the cortical p r o j e c t i o n o f LC ~s ~psdateral, by reason o f N A depletion m the Jpsilateral cortex following umlateral destruction o f LC2,3,7,17. K o r f et al. 'q a n d W a l t e r a n d Eccleston t8 s u p p o r t e d the t h e o r y o f ipsdateral cortical mnervation. They considered slight changes o f N A and ~ts m e t a b o h t e s m the c o n t r a l a t e r a l cortex as p r o b a b l y due to m & r e c t s t i m u l a t i o n as a result o f spread o f c u r r e n t from the electrode. Shinuzu et al. xe', however, d e m o n s t r a t e d w~th their m o r p h o l o g i c a l ewdence that the u n d a t e r a l lesion to L C produces a considerable n u m b e r o f d e g e n e r a t e d fibers n o t only in the ipsdateral cortex b u t also m the c o n t r a lateral cortex. Rets and Ross a5 have also f o u n d that d o p a m m e - f l - h y d r o x y l a s e act w i t y in the rat r e m a i n e d lowered in the ipsilateral cortex for up to 45 days, and also fell transiently for 7 days m the c o n t r a l a t e r a l cortex following LC destruction. The present w o r k revealed t h a t [3H]NA release from the c o n t r a l a t e r a l cortex occurred following stimulation o f the ventral p a r t o f LC a n d the region rostral a n d medml to LC. O u r findings that s t l m u l a h o n o f the LC results m N A release not only from the lpsdateral cortex but also f r o m the c o n t r a l a t e r a l cortex, are considered to s u p p o r t the hypothesis o f a bilateral cortical innervation from the LC. F u r t h e r experiments, e.g. cutting o f the c o n t r a l a t e r a l d o r s a l N A bundle, are necessary to rule out direct or indirect increase m ~mpulse flow t h r o u g h the c o n t r a l a t e r a l LC. This w o r k was s u p p o r t e d by g r a n t N o . 944041 f r o m the Ministry o f E d u c a h o n , Japan. The a u t h o r s express their gratitude to Prof. S. T a k a o r i for p e r t i n e n t advice a n d to M. O h a r a for assistance with the manuscript.
1 ANDI~N,N.-E., DAHLSTROM,A., FUXE, K , OLSON, L., AND UNGERSTEDT,U., Ascending monoamine neurons to the telencephalon and &encephalon, .4cta physiol, scand., 67 (1966) 313-326 2 ANLEZARK,G., CROW, J., AND GREENWAY,A., Impaired learning and decreased corhcal norepinephrine after bdateral locus coeruleus lesions, Scwnce, 181 (1973) 682-684. 3 ARBUTHNOTT,G W , CROW, T. J., FuxE, K., OLSON, L., AND UNGERSTEDT,U., Depletion of catecholamines in vivo induced by electrical stimulation of central monoamine pathways, Brain Research, 24 (1970) 471-483. 4 BRENELLS,A. B., Spontaneous and neurally evoked release of labelled noradrenahne from rabbit olfactory bulbs in vivo, J. Physiol. (Lond.), 240 (1974) 279-293 5 GADDUM,J. H., Push-pull cannulae, J Physiol. (Lond.), 155 (1961) 1-2. 6 FALCK,B., HILLARP,N -A., THIEME,G., ANDTOR, A., Fluorescence of catecholammes and related compounds condensed wxth formaldehyde, J. Histochem. Cytochem., 10 (1962) 348-354. 7 KOBAYASHI,R. M., PALKOVlTS,M., KOPIN, I. J., AND JACOBOWlTZ,D. M., B~ochem~cal mapping of noradrenergic nerves arising from the rat locus coeruleus, Brain Research, 77 (1974) 264-269. 8 KORF,J , AGHAJANIAN,G. K , AND ROTH, R. H., Stimulation and destruction of the locus coeruleus opposite effects on 3-methoxy-4-hydroxyphenylglycol sulfate levels m the rat cerebral cortex, Europ. J. PharmacoL, 2l (1973) 305-310. 9 KORF,J., ROTH, R. H., AND AGHAJANIAN,G. K , Alterations m turnover and endogenous levels of norepinephrine m cerebral cortex following electrical stimulation and acute axotomy of cerebral noradrenergic pathways, Europ. J. Pharmacol., 23 (1973) 276-282.
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