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Effects of selective toxic lesions of cholinergic neurons of the laterodorsal tegmental nucleus on experimental seizures
Brain Research, 579 (1992) 161-164 © 1992 Elsevier Science Publishers B.V. All fights reserved. 0006-8993/92/$05.00
161
BRES 25141
Effects of selective toxic lesions of cholinergic neurons of the laterodorsal tegmental nucleus on experimental seizures John W. Miller, Beverly C. Gray and Mark E. Bardgett Department of Neurology and Neurological Surgery (Neurology), Washington University School of Medicine, St. Louis, MO 63110 (USA)
(Accepted 21 January 1992) Key words: Laterodorsal tegmental nucleus; Seizure; Pentylenetetrazol; Reticular formation; Central medical nucleus; Ethylcholine mustard aziridinium ion; Arousal
This study determined the effects of bilateral discrete partial lesions of cholinergic neurons of the laterodorsal tegmental nucleus (LDTg) of the pontomesencephalic tegmentum on seizures induced by intravenous pentylenetetrazol (PTZ). Relatively selective lesions produced by bilateral 50 nl microinjections of 75 pmol of the cholinergic neurotoxin ethylcholine mustard aziridinium ion (AF64A) resulted in a significant reduction in the threshold of myoclonic and facial-forelimb clonic seizures but not tonic seizures when PTZ was infused 7 days later. This demonstrates that this cholinergic nucleus is a key site of subcortical seizure regulation. We propose that this control is mediated by ascending projections from the LDTg to the central medial intralaminar nucleus of the thalamus. Epileptic seizures often occur preferentially during drowsiness and slow wave sleep in humans and experimental animals 6'1s'19. This suggests that the neuroanatomical systems which mediate arousal also regulate seizures, although until recently the responsible nuclei were undefined. We have shown that discrete microinjections of y-aminobutyric acid (GABA) agonists in the laterodorsal tegmental nucleus (LDTg) of the pontomesencephalic tegmentum, depress arousal and facilitate a variety of seizure types while injections in nearby cholinergic and non-cholinergic nuclei do not 11. The purpose of the present study is to further investigate the role of the LDTg in seizure control by determining the elfects of destruction of its cholinergic neurons on seizures induced by intravenous pentylenetetrazol (PTZ). This was undertaken, in part, to confirm that the LDTg, rather than adjacent structures, regulates seizures, and to test the hypothesis that it is the cholinergic neurons of this nucleus which control seizure threshold. In addition, while our prior studies H have only measured seizure thresholds in the acute state induced by local injection of inhibitory agents, we now wanted to see if effects persist in a chronic model, Lesions were produced in the LDTg by bilateral 50 nl microinjections of the selective cholinergic neurotoxln ethylcholine mustard aziridinium ion (AF64A) 7 or control injections of 2% Alcian blue dye in normal saline, A F 6 4 A was prepared by maintaining a solution of acetylethylcholine mustard hydrochloride (Research Bio-
chemicals Inc.) at pH 11.3-11.7 for 30 min at room temperature, and then adding Alcian blue and adjusting the p H to 7.4 prior to use 3. Injections were made over 2 min with a 0.5/A Hamilton syringe in female Sprague-Dawley rats (150-180 g, Sasco) under halothane anesthesia (2-3% in 100% 02) at stereotaxic coordinates from the atlas of Paxinos and Watson 14. Six days later, jugular vein catheters were inserted and sutured under the skin using halothane anesthesia. The following day, the catheters were flushed with heparin and connected to a pump for continuous infusion of PTZ (Sigma, 5.0 mg/ml at a rate of 1.44 ml/min). This initially resulted in myoclonic jerks of the face, upper limbs and sometimes the entire body. This was followed by clonic seizure episodes ('facial-forelimb clonus '1) consisting of clonic forelimb movement, writhing trunk movements and intermixed facial or body myoclonic movements. Eventually a tonic seizure followed, consisting of trunk stiffening and tonic forelimb extension without hindlimb extension. The threshold for each seizure type was defined as the dose of convulsant per kg, calculated from the time of occurrence of the first seizure of that kind. It has been shown with this technique that the threshold doses and resultant brain P T Z levels are independent of infusion rate 13'15 implying rapid equilibration of the infused agent with its site of action. After the tonic seizure, the animals were sacrificed with 175 mg/kg intravenous sodium pentobarbital. The brains were removed, frozen, sectioned on a cryostat at 32/~m and stained to evaluate the locations
Correspondence: J.W. Miller, Department of Neurology, Box 8111, Washington University School of Medicine, 660 S. Euclid, St. Louis, MO 63110, USA. Fax: (1) (314) 362-2826.
163 TABLE I Group
n
PTZ threshold (mg/kg) Myoclonic
Control Lesion LDTg Misses
17
26.0 _+ 3.1
15 21
15.1 _+ 1.9 23.1 _+ 2.7
q
aT
Clonic
q
aT
31.5 + 3.7 3.96 1.29
<0.01 NS
19.0 _ 2.4 27.2 _+ 3.2
Tonic
q
aT
1.31 1.16
NS NS
80.6 _+ 9.1 3.60 1.34
<0.05 NS
68.8 + 8.1 70.9 _+ 7.7
Comparison of pentylenetetrazol (PTZ) seizure thresholds for myoclonic, facial-forelimb clonic (labeled as 'clonic') and tonic seizures between animals with bilateral control injections of 50 nl of 2% Alcian blue-normal saline and animals with lesions made with 75 pmol of AF64A in 50 nl. The group labeled 'LDTg' consists of animals with partial bilateral LDTg damage, excluding those with injections centered outside the LDTg, or significant damage to non-cholinergic neurons or to the cholinergic neurons of the pedunculopontine tegmental nucleus. The group labeled 'Misses' consists of animals where bilateral AF64A injections were misplaced, causing no apparent loss of NADPH-diaphorase staining neurons of the LDTg. Although the injections in this group had a variety of locations, most were ventral to the LDTg. The Student-Newman-Keuls test was used for statistical analysis, with q and a T values calculated relative to the control group.
and effects of the injections. The LDTg was identified by NADPH-diaphorase histochemistry 17, which selectively stains cholinergic neurons in this nucleus, as well as in the pedunculopontine tegmental nucleus ventrolateral to it 2° (Fig. 1). While diaphorase-staining neurons were not reduced with unilateral or bilateral injections of 10 or 25 pmol of AF64A, injections of 500 pmol caused local non-selective loss of all neural elements (Fig. 1). Injections of 75 or 100 pmol led to a partial loss of diaphorase staining in the LDTg without apparent damage to non-cholinergic neurons or to nearby cholinergic neurons of the pedunculopontine tegmental nucleus (Fig. 1). This narrow dose range for specific cholinotoxic effects is similar to that seen with injections of the nucleus basalis 7. Accordingly, only bilateral injections of 75 pmol of AF64A were used in studies of effects on seizure thresholds. There was no obvious change in arousal state or any other aspect of spontaneous behavior in these lesioned animals, either after they awoke from anesthesia, or prior to seizure induction. Only control animals with injections centered within the LDTg bilaterally (51% of controls), or experimental animals with either bilateral partial cell loss within this nucleus without damage to non-cholinergic neurons or to cholinergic neurons of the pedunculopontine tegmental nucleus (31% of animals with AF64A injections) or with complete sparing of the
LDTg due to misplaced injections were included in the data analysis (Table I). In the animals with bilateral LDTg damage there was a significant decrease in the thresholds of myoclonic and facial forelimb clonic seizures, without a significant effect on tonic seizures (Table I). There was no change in the appearance of the lesioned animals' ictal behavior during any of these seizure types compared to controls. Lesions outside this nucleus did not significantly affect seizures. The main finding of this study, therefore, is that very small, selective partial lesions of the LDTg produce clear changes in PTZ seizure thresholds which are identical to, though smaller in magnitude than, those produced by LDTg microinjections of the GABA A agonist, piperidine-4-sulfonic acid, or the GABA B agonist, (-)baclofen 11. This is consistent with the hypothesis that cholinergic neurons of this nucleus control susceptibility to seizures. In contrast, these lesions did not duplicate effects on spontaneous behavior previously observed after LDTg microinjections of GABA agonists 11. Such injections produced a hypoactive condition with the animals lying limply on their sides, unable to right, without spontaneous movement except respirations. The LDTg mediates these effects, since mjcroinjections in surrounding nuclei have little influence on behavior. The lack of behavioral effects of the lesions cannot be attributed to the selec-
Fig. 1. Photomicrographs of the pontomesencephalic tegmentum showing cholinergic neurons demonstrated with NADPH-diaphorase stain in unfixed tissue. The top micrograph shows the normal anatomy of this region, with a dense cluster of cholinergic neurons in the periventricular gray matter forming the laterodorsal tegmental nucleus (LDTg), and neurons of the pedunculopontine tegmental nucleus Cstreaming out ventrolaterally. The micrograph in the center shows non-selective loss of all neural elements at the site (arrow) of a unilateral injection of 500 pmol of ethylcholine mustard aziridinium ion (AF64A). The micrograph on the bottom panel shows bilateral injections of 75 pmol of AF64A, with the arrows marking small clumps of Alcian blue dye at the injection sites. This micrograph demonstrates loss of NADPHdiaphorase staining neurons in the LDTg, much greater on the left than the right side, without involvement of nearby pedunculopontine tegmental neurons.
164 tivity of A F 6 4 A for cholinergic neurons, because electrolytic5 and excitotoxical lesions of the L D T g and surrounding pontomesencephalic t e g m e n t u m in the cat also do not produce obvious acute changes in behavior. R E M sleep without atonia is seen with such lesions 5'21, although this has been attributed to a region ventral to the L D T g and adjacent to the locus coeruleus TM. The disparity between behavioral effects of lesions and injections must be due to the p r o f o u n d differences between the physiological nature of these two experimental manipulations, since one consists of the p e r m a n e n t loss of neuronal cell bodies and their dendritic and axonal processes, p r e s u m a b l y with associated trophic changes in postsynaptic elements, and the other of sudden, selective, intense activation of local G A B A receptors. We have found similar differences in behavioral effects of G A B A injections and lesions in the central medial thalamic nucleus (references 8-11 and unpublished obser-
clonic and facial-forelimb clonic seizures. These seizure types originate in the forebrain 1°, with evidence for this including the fact that they are abolished by high brainstem transections ~. It is therefore likely that the L D T g controls seizures by means of ascending projections. We have previously shown that injections of G A B A agonists in the central medial intralaminar nucleus of the midline thalamus also depress arousal and facilitate myoclonic and clonic seizures in the same fashion as L D T g microinjections, while injections in surrounding thalamic structures have little effect 8-11. Microinjections of muscarinic antagonists in the central medial nucleus also depress arousal and seizure thresholds ~2. The important direct cholinergic projections from the L D T g to the central medial nucleus 4 and these effects of G A B A e r g i c and cholinergic agents, suggest that these two nuclei form an ascending, midline seizure regulating system.
vations). L D T g lesions and inhibitory injections affect only myo-
This work was supported by NIH Grants NS01296 and NS14834. We would like to thank Morgan Raley for his technical assistance.
1 Browning, R.A. and Nelson, D.K., Modification of electroshock and pentylenetetrazol seizure patterns in rats after precollicular transections, Exp. Neurol., 93 (1986) 546-556. 2 Culebras, A. and Moore, J.T., Magnetic resonance findings in REM sleep behavior disorder, Neurology, 39 (1989) 1519-1523. 3 Fisher, A., Mantione, C.R., Abraham, D.J. and Hanin, I., Long-term central cholinergic hypofunction induced in mice by ethylcholine aziridinium ion (AF64A) in vivo, J. Pharmacol. Exp. Ther., 222 (1982) 140-145. 4 Hallinger, A.E., Levey, A.I., Lee, H.J., Rye, D.B. and Wainer, B.H., The origins of cholinergic and other subcortical afferents to the thalamus in the rat, J. Comp. Neurol., 262 (1987) 105124. 5 Hendricks, J.C., Morrison, A.R. and Mann, G.L., Different behaviors during paradoxical sleep without atonia depend on pontine lesion site, Brain Res., 239 (1982) 81-105. 6 Klass, D.W. and Fischer-Williams, M., Sensory stimulation, sleep and sleep deprivation. In A. Remond (Ed.), Handbook of Electroencephalography and Clinical Neurophysiology, Vol. 13D, Elsevier, Amsterdam, 1973, pp. 5-73. -7 Kozlowski, M.R. and Arbogast, R.I., Specific toxic effects of ethylcholine nitrogen mustard on cholinergic neurons of the nucleus basalis of Meynert, Brain Res., 372 (1986) 45-54. 8 Miller, J.W., Hall, C.M., Holland, K. and Ferrendelli, J.A., Identification of a median thalamic system regulating seizures and arousal, Epilepsia, 30 (1989) 493-500. 9 Miller, J.W. and Ferrendelli, J.A, The central medial nucleus: thalamic site of seizure regulation, Brain Res., 508 (1990) 297300. 10 Miller, J.W. and Ferrendelli, J.A., Characterization of GABAergic seizure regulation in the midline thalamus, Neuropharmacology, 29 (1990) 649-655. 11 Miller, J.W., Bardgett, M.E. and Gray, B.C., The role of the laterodorsal tegmental nucleus of the rat in experimental seizures, Neuroscience, 29 (1991) 649-655. 12 Miller, J.W., Bardgett, M.E. and Gray, B.C., Characterization
of cholinergic mechanisms regulating seizures in the midline thalamus, Neuropharmacology, in press. 13 Nutt, D.J., Cowen, D.J. and Green, A.R., On the measurement in rats of the convulsant effect of drugs and the changes which follow electroconvulsive shock, Neuropharmacology, 19 (1980) 1017-1023. 14 Paxinos, G. and Watson, C., The Rat Brain in Stereotaxic Coordinates, Academic Press, New York, 1982. 15 Ramzan, I.M. and Levy, G., Kinetics of drug action in disease states. XIV. Effect of infusion rate on pentylenetetrazol concentrations in serum, brain and cerebrospinal fluid of rats at onset of convulsions, J. Pharmacol. Exp. Ther., 234 (1985) 624628. 16 Sakai, K., Sastre, J.P., Salvert, D., Touret, M., Tohyama, M. and Jouvet, M., Tegmentoreticular projections with special reference to the muscular atonia during paradoxical sleep in the cat: an HRP study, Brain Res., 176 (1979) 233-254. 17 Scherer-Singler; U., Vincent, S.R., Kimura, H. and McGeer, E.G., Demonstration of a unique population of neurons with NADPH-diaphorase histochemistry, J. Neurosci. Meth., 9 (1983) 229-234. 18 Shouse, M.N., Sleep deprivation increases susceptibility to kindied and penicillin seizure events during all waking and sleep states in cats, Sleep, 11 (1988) 162-171. 19 Shouse, M.N., Sleep deprivation increases thalamocortical excitability in the somatomotor pathway, especially during seizure-prone sleep or awakening states in feline seizure models, Exp. Neurol., 99 (1988) 664-667. 20 Vincent, S.R., Satoh, K., Armstrong, D.M., Panula, P., Vale, W. and Fibiger, H.C., Neuropeptides and NADPH-diaphorase activity in the ascending cholinergic reticular system of the rat, Neuroscience, 17 (1986) 167-182. 21 Webster, H.H. and Jones, B.E., Neurotoxic lesions of the dorsolateral pontomesencephalic tegmentum-cholinergic cell area in the cat. II. Effects upon sleep-waking states, Brain Res., 458 (1988) 285-302.
161
BRES 25141
Effects of selective toxic lesions of cholinergic neurons of the laterodorsal tegmental nucleus on experimental seizures John W. Miller, Beverly C. Gray and Mark E. Bardgett Department of Neurology and Neurological Surgery (Neurology), Washington University School of Medicine, St. Louis, MO 63110 (USA)
(Accepted 21 January 1992) Key words: Laterodorsal tegmental nucleus; Seizure; Pentylenetetrazol; Reticular formation; Central medical nucleus; Ethylcholine mustard aziridinium ion; Arousal
This study determined the effects of bilateral discrete partial lesions of cholinergic neurons of the laterodorsal tegmental nucleus (LDTg) of the pontomesencephalic tegmentum on seizures induced by intravenous pentylenetetrazol (PTZ). Relatively selective lesions produced by bilateral 50 nl microinjections of 75 pmol of the cholinergic neurotoxin ethylcholine mustard aziridinium ion (AF64A) resulted in a significant reduction in the threshold of myoclonic and facial-forelimb clonic seizures but not tonic seizures when PTZ was infused 7 days later. This demonstrates that this cholinergic nucleus is a key site of subcortical seizure regulation. We propose that this control is mediated by ascending projections from the LDTg to the central medial intralaminar nucleus of the thalamus. Epileptic seizures often occur preferentially during drowsiness and slow wave sleep in humans and experimental animals 6'1s'19. This suggests that the neuroanatomical systems which mediate arousal also regulate seizures, although until recently the responsible nuclei were undefined. We have shown that discrete microinjections of y-aminobutyric acid (GABA) agonists in the laterodorsal tegmental nucleus (LDTg) of the pontomesencephalic tegmentum, depress arousal and facilitate a variety of seizure types while injections in nearby cholinergic and non-cholinergic nuclei do not 11. The purpose of the present study is to further investigate the role of the LDTg in seizure control by determining the elfects of destruction of its cholinergic neurons on seizures induced by intravenous pentylenetetrazol (PTZ). This was undertaken, in part, to confirm that the LDTg, rather than adjacent structures, regulates seizures, and to test the hypothesis that it is the cholinergic neurons of this nucleus which control seizure threshold. In addition, while our prior studies H have only measured seizure thresholds in the acute state induced by local injection of inhibitory agents, we now wanted to see if effects persist in a chronic model, Lesions were produced in the LDTg by bilateral 50 nl microinjections of the selective cholinergic neurotoxln ethylcholine mustard aziridinium ion (AF64A) 7 or control injections of 2% Alcian blue dye in normal saline, A F 6 4 A was prepared by maintaining a solution of acetylethylcholine mustard hydrochloride (Research Bio-
chemicals Inc.) at pH 11.3-11.7 for 30 min at room temperature, and then adding Alcian blue and adjusting the p H to 7.4 prior to use 3. Injections were made over 2 min with a 0.5/A Hamilton syringe in female Sprague-Dawley rats (150-180 g, Sasco) under halothane anesthesia (2-3% in 100% 02) at stereotaxic coordinates from the atlas of Paxinos and Watson 14. Six days later, jugular vein catheters were inserted and sutured under the skin using halothane anesthesia. The following day, the catheters were flushed with heparin and connected to a pump for continuous infusion of PTZ (Sigma, 5.0 mg/ml at a rate of 1.44 ml/min). This initially resulted in myoclonic jerks of the face, upper limbs and sometimes the entire body. This was followed by clonic seizure episodes ('facial-forelimb clonus '1) consisting of clonic forelimb movement, writhing trunk movements and intermixed facial or body myoclonic movements. Eventually a tonic seizure followed, consisting of trunk stiffening and tonic forelimb extension without hindlimb extension. The threshold for each seizure type was defined as the dose of convulsant per kg, calculated from the time of occurrence of the first seizure of that kind. It has been shown with this technique that the threshold doses and resultant brain P T Z levels are independent of infusion rate 13'15 implying rapid equilibration of the infused agent with its site of action. After the tonic seizure, the animals were sacrificed with 175 mg/kg intravenous sodium pentobarbital. The brains were removed, frozen, sectioned on a cryostat at 32/~m and stained to evaluate the locations
Correspondence: J.W. Miller, Department of Neurology, Box 8111, Washington University School of Medicine, 660 S. Euclid, St. Louis, MO 63110, USA. Fax: (1) (314) 362-2826.
163 TABLE I Group
n
PTZ threshold (mg/kg) Myoclonic
Control Lesion LDTg Misses
17
26.0 _+ 3.1
15 21
15.1 _+ 1.9 23.1 _+ 2.7
q
aT
Clonic
q
aT
31.5 + 3.7 3.96 1.29
<0.01 NS
19.0 _ 2.4 27.2 _+ 3.2
Tonic
q
aT
1.31 1.16
NS NS
80.6 _+ 9.1 3.60 1.34
<0.05 NS
68.8 + 8.1 70.9 _+ 7.7
Comparison of pentylenetetrazol (PTZ) seizure thresholds for myoclonic, facial-forelimb clonic (labeled as 'clonic') and tonic seizures between animals with bilateral control injections of 50 nl of 2% Alcian blue-normal saline and animals with lesions made with 75 pmol of AF64A in 50 nl. The group labeled 'LDTg' consists of animals with partial bilateral LDTg damage, excluding those with injections centered outside the LDTg, or significant damage to non-cholinergic neurons or to the cholinergic neurons of the pedunculopontine tegmental nucleus. The group labeled 'Misses' consists of animals where bilateral AF64A injections were misplaced, causing no apparent loss of NADPH-diaphorase staining neurons of the LDTg. Although the injections in this group had a variety of locations, most were ventral to the LDTg. The Student-Newman-Keuls test was used for statistical analysis, with q and a T values calculated relative to the control group.
and effects of the injections. The LDTg was identified by NADPH-diaphorase histochemistry 17, which selectively stains cholinergic neurons in this nucleus, as well as in the pedunculopontine tegmental nucleus ventrolateral to it 2° (Fig. 1). While diaphorase-staining neurons were not reduced with unilateral or bilateral injections of 10 or 25 pmol of AF64A, injections of 500 pmol caused local non-selective loss of all neural elements (Fig. 1). Injections of 75 or 100 pmol led to a partial loss of diaphorase staining in the LDTg without apparent damage to non-cholinergic neurons or to nearby cholinergic neurons of the pedunculopontine tegmental nucleus (Fig. 1). This narrow dose range for specific cholinotoxic effects is similar to that seen with injections of the nucleus basalis 7. Accordingly, only bilateral injections of 75 pmol of AF64A were used in studies of effects on seizure thresholds. There was no obvious change in arousal state or any other aspect of spontaneous behavior in these lesioned animals, either after they awoke from anesthesia, or prior to seizure induction. Only control animals with injections centered within the LDTg bilaterally (51% of controls), or experimental animals with either bilateral partial cell loss within this nucleus without damage to non-cholinergic neurons or to cholinergic neurons of the pedunculopontine tegmental nucleus (31% of animals with AF64A injections) or with complete sparing of the
LDTg due to misplaced injections were included in the data analysis (Table I). In the animals with bilateral LDTg damage there was a significant decrease in the thresholds of myoclonic and facial forelimb clonic seizures, without a significant effect on tonic seizures (Table I). There was no change in the appearance of the lesioned animals' ictal behavior during any of these seizure types compared to controls. Lesions outside this nucleus did not significantly affect seizures. The main finding of this study, therefore, is that very small, selective partial lesions of the LDTg produce clear changes in PTZ seizure thresholds which are identical to, though smaller in magnitude than, those produced by LDTg microinjections of the GABA A agonist, piperidine-4-sulfonic acid, or the GABA B agonist, (-)baclofen 11. This is consistent with the hypothesis that cholinergic neurons of this nucleus control susceptibility to seizures. In contrast, these lesions did not duplicate effects on spontaneous behavior previously observed after LDTg microinjections of GABA agonists 11. Such injections produced a hypoactive condition with the animals lying limply on their sides, unable to right, without spontaneous movement except respirations. The LDTg mediates these effects, since mjcroinjections in surrounding nuclei have little influence on behavior. The lack of behavioral effects of the lesions cannot be attributed to the selec-
Fig. 1. Photomicrographs of the pontomesencephalic tegmentum showing cholinergic neurons demonstrated with NADPH-diaphorase stain in unfixed tissue. The top micrograph shows the normal anatomy of this region, with a dense cluster of cholinergic neurons in the periventricular gray matter forming the laterodorsal tegmental nucleus (LDTg), and neurons of the pedunculopontine tegmental nucleus Cstreaming out ventrolaterally. The micrograph in the center shows non-selective loss of all neural elements at the site (arrow) of a unilateral injection of 500 pmol of ethylcholine mustard aziridinium ion (AF64A). The micrograph on the bottom panel shows bilateral injections of 75 pmol of AF64A, with the arrows marking small clumps of Alcian blue dye at the injection sites. This micrograph demonstrates loss of NADPHdiaphorase staining neurons in the LDTg, much greater on the left than the right side, without involvement of nearby pedunculopontine tegmental neurons.
164 tivity of A F 6 4 A for cholinergic neurons, because electrolytic5 and excitotoxical lesions of the L D T g and surrounding pontomesencephalic t e g m e n t u m in the cat also do not produce obvious acute changes in behavior. R E M sleep without atonia is seen with such lesions 5'21, although this has been attributed to a region ventral to the L D T g and adjacent to the locus coeruleus TM. The disparity between behavioral effects of lesions and injections must be due to the p r o f o u n d differences between the physiological nature of these two experimental manipulations, since one consists of the p e r m a n e n t loss of neuronal cell bodies and their dendritic and axonal processes, p r e s u m a b l y with associated trophic changes in postsynaptic elements, and the other of sudden, selective, intense activation of local G A B A receptors. We have found similar differences in behavioral effects of G A B A injections and lesions in the central medial thalamic nucleus (references 8-11 and unpublished obser-
clonic and facial-forelimb clonic seizures. These seizure types originate in the forebrain 1°, with evidence for this including the fact that they are abolished by high brainstem transections ~. It is therefore likely that the L D T g controls seizures by means of ascending projections. We have previously shown that injections of G A B A agonists in the central medial intralaminar nucleus of the midline thalamus also depress arousal and facilitate myoclonic and clonic seizures in the same fashion as L D T g microinjections, while injections in surrounding thalamic structures have little effect 8-11. Microinjections of muscarinic antagonists in the central medial nucleus also depress arousal and seizure thresholds ~2. The important direct cholinergic projections from the L D T g to the central medial nucleus 4 and these effects of G A B A e r g i c and cholinergic agents, suggest that these two nuclei form an ascending, midline seizure regulating system.
vations). L D T g lesions and inhibitory injections affect only myo-
This work was supported by NIH Grants NS01296 and NS14834. We would like to thank Morgan Raley for his technical assistance.
1 Browning, R.A. and Nelson, D.K., Modification of electroshock and pentylenetetrazol seizure patterns in rats after precollicular transections, Exp. Neurol., 93 (1986) 546-556. 2 Culebras, A. and Moore, J.T., Magnetic resonance findings in REM sleep behavior disorder, Neurology, 39 (1989) 1519-1523. 3 Fisher, A., Mantione, C.R., Abraham, D.J. and Hanin, I., Long-term central cholinergic hypofunction induced in mice by ethylcholine aziridinium ion (AF64A) in vivo, J. Pharmacol. Exp. Ther., 222 (1982) 140-145. 4 Hallinger, A.E., Levey, A.I., Lee, H.J., Rye, D.B. and Wainer, B.H., The origins of cholinergic and other subcortical afferents to the thalamus in the rat, J. Comp. Neurol., 262 (1987) 105124. 5 Hendricks, J.C., Morrison, A.R. and Mann, G.L., Different behaviors during paradoxical sleep without atonia depend on pontine lesion site, Brain Res., 239 (1982) 81-105. 6 Klass, D.W. and Fischer-Williams, M., Sensory stimulation, sleep and sleep deprivation. In A. Remond (Ed.), Handbook of Electroencephalography and Clinical Neurophysiology, Vol. 13D, Elsevier, Amsterdam, 1973, pp. 5-73. -7 Kozlowski, M.R. and Arbogast, R.I., Specific toxic effects of ethylcholine nitrogen mustard on cholinergic neurons of the nucleus basalis of Meynert, Brain Res., 372 (1986) 45-54. 8 Miller, J.W., Hall, C.M., Holland, K. and Ferrendelli, J.A., Identification of a median thalamic system regulating seizures and arousal, Epilepsia, 30 (1989) 493-500. 9 Miller, J.W. and Ferrendelli, J.A, The central medial nucleus: thalamic site of seizure regulation, Brain Res., 508 (1990) 297300. 10 Miller, J.W. and Ferrendelli, J.A., Characterization of GABAergic seizure regulation in the midline thalamus, Neuropharmacology, 29 (1990) 649-655. 11 Miller, J.W., Bardgett, M.E. and Gray, B.C., The role of the laterodorsal tegmental nucleus of the rat in experimental seizures, Neuroscience, 29 (1991) 649-655. 12 Miller, J.W., Bardgett, M.E. and Gray, B.C., Characterization
of cholinergic mechanisms regulating seizures in the midline thalamus, Neuropharmacology, in press. 13 Nutt, D.J., Cowen, D.J. and Green, A.R., On the measurement in rats of the convulsant effect of drugs and the changes which follow electroconvulsive shock, Neuropharmacology, 19 (1980) 1017-1023. 14 Paxinos, G. and Watson, C., The Rat Brain in Stereotaxic Coordinates, Academic Press, New York, 1982. 15 Ramzan, I.M. and Levy, G., Kinetics of drug action in disease states. XIV. Effect of infusion rate on pentylenetetrazol concentrations in serum, brain and cerebrospinal fluid of rats at onset of convulsions, J. Pharmacol. Exp. Ther., 234 (1985) 624628. 16 Sakai, K., Sastre, J.P., Salvert, D., Touret, M., Tohyama, M. and Jouvet, M., Tegmentoreticular projections with special reference to the muscular atonia during paradoxical sleep in the cat: an HRP study, Brain Res., 176 (1979) 233-254. 17 Scherer-Singler; U., Vincent, S.R., Kimura, H. and McGeer, E.G., Demonstration of a unique population of neurons with NADPH-diaphorase histochemistry, J. Neurosci. Meth., 9 (1983) 229-234. 18 Shouse, M.N., Sleep deprivation increases susceptibility to kindied and penicillin seizure events during all waking and sleep states in cats, Sleep, 11 (1988) 162-171. 19 Shouse, M.N., Sleep deprivation increases thalamocortical excitability in the somatomotor pathway, especially during seizure-prone sleep or awakening states in feline seizure models, Exp. Neurol., 99 (1988) 664-667. 20 Vincent, S.R., Satoh, K., Armstrong, D.M., Panula, P., Vale, W. and Fibiger, H.C., Neuropeptides and NADPH-diaphorase activity in the ascending cholinergic reticular system of the rat, Neuroscience, 17 (1986) 167-182. 21 Webster, H.H. and Jones, B.E., Neurotoxic lesions of the dorsolateral pontomesencephalic tegmentum-cholinergic cell area in the cat. II. Effects upon sleep-waking states, Brain Res., 458 (1988) 285-302.