Gene targeting reveals a critical role for neurturin in the development and maintenance of enteric, sensory, and parasympathetic neurons.

Neurturin (NTN) is a neuronal survival factor that activates the Ret tyrosine kinase in the presence of a GPI-linked coreceptor (either GFR alpha1 or GFR alpha2). Neurturin-deficient (NTN-/-) mice generated by homologous recombination are viable and fertile but have defects in the enteric nervous system, including reduced myenteric plexus innervation density and reduced gastrointestinal motility. Parasympathetic innervation of the lacrimal and submandibular salivary gland is dramatically reduced in NTN-/- mice, indicating that Neurturin is a neurotrophic factor for parasympathetic neurons. GFR alpha2-expressing cells in the trigeminal and dorsal root ganglia are also depleted in NTN-/- mice. The loss of GFR alpha2-expressing neurons, in conjunction with earlier studies, provides strong support for GFR alpha2/Ret receptor complexes as the critical mediators of NTN function in vivo.

Progressive neurodegeneration after intracerebroventricular kainic acid administration in rats: implications for schizophrenia?

BACKGROUND: Intracerebroventricular (ICV) administration of kainic acid to rats produces limbic-cortical neuronal damage that has been compared to the neuropathology of schizophrenia. METHODS: Groups of adult rats were administered ICV kainic acid and then assessed for neuronal loss and the expression of proteins relevant to mechanisms of neuronal damage after one and fourteen days. Neuronal loss was assessed by two-dimensional cell counting and protein expression was assessed by immunohistochemistry. RESULTS: ICV kainic acid administration was associated with both immediate (day 1) and delayed (day 14) neuronal loss in the dorsal hippocampus. The immediate injury was largely limited to the CA3 hippocampal subfield, while the delayed injury included the CA1 subfield. Multiple mechanisms of cell death appeared to be involved in the delayed neuronal loss, as evidenced by changes in the expression of glutamate receptor subunits, heat shock protein and jun protein. CONCLUSIONS: ICV kainic acid administration to adult rats produces progressive damage to limbic-cortical neurons, involving both fast and slow mechanisms of cell death. Given the evidence for clinical deterioration, cognitive deficits and hippocampal neuropathy in some cases of schizophrenia, this animal model may be relevant for hypotheses regarding mechanisms of neurodegeneration in that disorder.

Platelet serotonin markers and depressive symptomatology.

Dysfunction of brain serotonergic symptoms may be a factor in the mood and behavioral disturbances associated with depression. Platelet serotonin measures represent indirect but easily obtainable indices of brain serotonin function. To examine the specificity of relationships between cognitive and vegetative symptom groupings and platelet serotonin measures, we assessed 35 depressed outpatients using the Hamilton Rating Scale for Depression and collected platelets after a minimum 3-week drug-free period. Platelets were also collected from 14 controls. The results showed that depressed patients had lower platelet serotonin (5-HT) uptake site density values than controls and that 5-HT uptake site density values were inversely correlated with the severity of cognitive symptoms of depression. Platelet 5-HT2 receptor density values were higher in depressed patients than controls, and there was a trend toward a direct correlation between the cognitive symptoms of depression and 5-HT2 receptor density values. Neither platelet measure showed any relationship with the severity of the vegetative symptoms of depression.

CSF excitatory amino acids and severity of illness in Alzheimer's disease.

Researchers have proposed that increased release of excitatory amino acids (EAAs) is involved in the pathogenesis of dementia of the Alzheimer type (DAT), and CSF EAA concentrations have been measured to obtain evidence in support of this hypothesis. However, previous comparisons of CSF EAA concentrations in patients with DAT and in controls have yielded inconsistent results, perhaps because patient samples have been heterogeneous as to dementia severity. To determine whether there are changes in CSF concentrations of EAAs related to severity of illness in patients with DAT, we measured CSF concentrations of glutamate, aspartate, and taurine in 32 subjects with DAT, in whom we also assessed the severity of illness using clinical and neuropsychological measures, and 11 age-matched controls. The results suggested that increased CSF aspartate and glutamate concentrations, as well as decreased taurine concentrations, may occur in some persons with more advanced symptoms of DAT.

Platelet binding characteristics distinguish placebo responders from nonresponders in depression.

To determine whether there are characteristics distinguishing placebo responders from nonresponders, we studied 37 outpatients meeting DSM-III-R criteria for depression who were enrolled in controlled drug trials and 14 control subjects. Clinical data and blood samples were collected on admission and after a 7- to 10-day placebo washout. All patients experiencing a 40% drop in the Hamilton Rating Scale for Depression (HRSD) at the time of the second evaluation were considered placebo responders. There were no statistically significant differences between the two groups in clinical variables. Platelet markers distinguished the groups: Most notably, placebo nonresponders had the lowest 5-HT uptake site density values, placebo responders had intermediate values, and normal controls had the highest values. Placebo responders and placebo nonresponders had higher 5-HT uptake affinity values. No significant differences were observed among the groups in platelet 5-HT2 receptor site density or affinity values. These results suggest that platelet serotonin characteristics, but not common clinical characteristics, may distinguish depressed patients who do and do not respond to placebo.

Characterization of cholinergic regulation of seizures by the midline thalamus.

This study determined the effects of injections of different cholinergic agents in the central medial intralaminar nucleus of the thalamus on seizures induced by intravenous injection of pentylenetetrazol. Injections of the cholinesterase inhibitor, neostigmine bromide, induced a stiff, tremulous state and lowered myoclonic, clonic and tonic seizure thresholds. The nicotinic agonist, tartrate, depressed arousal and facilitated all types of seizure, while its antagonist, d-tubocurarine chloride, heightened arousal and transformed pentylenetetrazol-induced convulsions, with tonic seizures occurring at a very low threshold without preceding myoclonic or clonic seizures or EEG spikes. The muscarinic agonist (+/-)pilocarpine hydrochloride, in very large doses, induced slight hyperactivity and facilitated tonic seizures but did not affect myoclonic or clonic seizures. Its antagonist, (-)scopolamine hydrobromide, slightly depressed arousal and myoclonic and clonic seizure thresholds. Injections of mixtures of agonists and antagonists (d-tubocurarine chloride + nicotine tartrate or (+/-)pilocarpine hydrochloride + (-)scopolamine) had little effect on spontaneous behavior or seizures. These results suggest that the midline thalamus regulates seizures and arousal, under the control of cholinergic neurotransmission. Nicotinic and muscarinic receptors have opposing roles in mediating these functions.

Locomotor activity and accumbens Fos expression driven by ventral hippocampal stimulation require D1 and D2 receptors.

Numerous studies have suggested that excitatory projections from the ventral hippocampus to the nucleus accumbens modulate locomotor activity in rats. Furthermore, the ability of ventral hippocampal neurons to alter locomotor activity may involve the dense dopaminergic innervation found in the nucleus accumbens. The purpose of this study was to: (i) more fully characterize the locomotor effects of acute alterations in ventral hippocampal activity; (ii) ascertain the influence of dopamine agonists and antagonists on locomotor changes produced by altered ventral hippocampal activity; and (iii) use immediate early gene induction to determine whether dopamine antagonists alter the response of nucleus accumbens neurons to ventral hippocampal stimulation. By comparing a variety of excitatory amino acid agonists, it was found that ventral hippocampal infusion of N-methyl-D-aspartate elevated locomotor activity in a subconvulsive manner, while other excitatory amino acid receptor agonists did not. Inactivation of the ventral hippocampus achieved by lidocaine infusion did not suppress ongoing locomotor activity, nor did it affect amphetamine-induced increases in locomotor activity. Increases in locomotor activity induced by ventral hippocampal N-methyl-D-aspartate infusion were blocked by systemic administration of haloperidol (a D2 receptor antagonist), SCH-23390 (a D1 receptor antagonist) or reserpine. Cellular expression of the protein product of the immediate early gene, c-fos, was dramatically increased in the nucleus accumbens shell after ventral hippocampal N-methyl-D-aspartate infusion, and haloperidol, SCH-23390 and reserpine attenuated this effect. These results suggest that the increases, but not decreases, in ventral hippocampal activity have a measurable effect on ongoing rates of locomotion, and that this effect requires both D1 and D2 receptors. Moreover, the studies of Fos expression suggest that dopamine receptor antagonists attenuate neuronal responses to ventral hippocampal stimulation within the nucleus accumbens, a brain region important in the generation and maintenance of locomotor activity.

The role of the laterodorsal tegmental nucleus of the rat in experimental seizures.

This study determined the effects of discrete microinjections of GABA agonists in the cholinergic nuclei of the pontomesencephalic tegmentum on spontaneous behavior and seizures induced by intravenous pentylenetetrazol, bicuculline or strychnine, in the rat. Injections of both the GABAA agonist piperidine-4-sulfonic acid and the GABAB agonist (-)baclofen in the laterodorsal tegmental nucleus produced a dose-dependent suppression of behavioral arousal and a reduction in the threshold of myoclonic and clonic but not tonic seizures induced by bicuculline and pentylenetetrazol. There were no significant effects on any type of strychnine seizure. Injections in the surrounding brainstem structures, including the pedunculopontine tegmental nucleus, had little effect on spontaneous behavior and did not significantly alter the thresholds of pentylenetetrazol-induced seizures. We have previously demonstrated that injections of GABA agonists in the central medial intralaminar nucleus of the thalamus have similar effects on behavior and seizures. Since the central medial nucleus receives important direct cholinergic projections from the laterodorsal tegmental nucleus, these two nuclei form a discrete ascending system which regulates seizure threshold.

Deletion of the N-terminus of murine map2 by gene targeting disrupts hippocampal ca1 neuron architecture and alters contextual memory.

Microtubule-associated protein-2 (MAP2) is a brain specific A-kinase anchoring protein that targets the cyclic AMP-dependent protein kinase holoenzyme (PKA) to microtubules. Phosphorylation of MAP2 by different protein kinases is crucial for neuronal growth. The N-terminus of MAP2 contains the binding site for regulatory subunit II of cAMP-dependent protein kinase (PKA-RIIbeta). Using homologous recombination, we created a mutant line of mice (delta1-158) that express truncated MAP2 lacking the N-terminal peptide and the PKA binding site. Deletion of the PKA binding site from the MAP2 gene resulted in decreased efficiency of MAP2 phosphorylation. Biochemical and immunohistochemical studies demonstrate major changes in the morphology of hippocampal neurons in delta1-158 mice. Behavioral tests indicate that delta1-158 mice were impaired (exhibited less conditioned freezing) relative to Wild-Type (WT) controls during a test of contextual, but not during auditory cue, fear conditioning when tested at 8 weeks or 8 months of age. The delta1-158 mice displayed a heightened sensitivity to shock at 8 weeks, but not at 8 months of age. We conclude that PKA binding to MAP2 and MAP2 phosphorylation is essential for the selective development of contextual memory.

Glucocorticoid interactions with memory function in schizophrenia.

Glucocorticoid (GC) exposure can affect brain function, including potential adverse effects on hippocampal physiology and on specific elements of cognitive performance. In a prior study of healthy adult humans, decreased verbal memory performance was detected during four days of double-blind, placebo-controlled dexamethasone (DEX) treatment. Using an identical experimental design and sample size (n = 19), the cognitive effect of DEX treatment was studied in 11 subjects with schizophrenia, compared with 8 receiving placebo. In contrast to the effect in healthy adults, GC treatment with DEX at this dose (cumulative 3.5 mg) and duration did not decrease verbal memory performance or other measures of cognitive function in the patients with schizophrenia. When data from this experiment was compared with data from the previous study of healthy adults, covarying differences in baseline memory performance, a significant 3-way interaction was detected between subject group, treatment condition, and the repeated measurements of verbal memory performance across baseline, treatment and washout (F[3,87] = 4.84, p = .0066), suggesting differential cognitive effects of DEX in the patients versus the previously studied healthy subjects. Baseline plasma cortisol concentrations (0800 h) prior to DEX treatment were inversely correlated with baseline delayed (rs = -0.536, p = .03) verbal recall performance, supporting a previous report. The current results await replication using a larger sample size but provide preliminary evidence for an altered behavioral response to acute GC exposure in schizophrenic versus healthy subjects, and further evidence for a relationship between chronic changes in circulating cortisol and the memory impairments found in this disorder.

Subcortical dopamine and serotonin turnover during acute and subchronic administration of typical and atypical neuroleptics.

The effects of acute (1 day) and subchronic (28 days) treatment with three atypical antipsychotic drugs [clozapine, (+/-)-sulpiride and (-)-3-PPP] on dopamine and serotonin turnover in both the nucleus accumbens (NA) and corpus striatum (CS) of rodents was compared to haloperidol and saline treatment. The equivalent doses of all drugs were determined based upon their ability to compete in vivo for 3H-spiperone binding in the NA and CS. All three atypical drugs, compared to haloperidol, produced preferential elevations of dopamine turnover in the NA. Further, the development of tolerance of this effect was more apparent for the three atypical drugs than for haloperidol. Surprisingly, all three atypical drugs, but not haloperidol, produced changes in serotonin turnover, despite the fact that (+/-)-sulpiride and (-)-3-PPP have no known direct effects on brain serotonin systems. All three atypical drugs produced acute increases in serotonin turnover in both the NA and CS, followed by later diseases.

The effects of kainic acid lesions on dopaminergic responses to haloperidol and clozapine.

The antipsychotic drugs haloperidol and clozapine have the common action of increasing dopamine metabolism in the striatum (nucleus accumbens, caudate-putamen) of the rat. Intracerebroventricular administration of kainic acid (KA) produces neuronal loss in limbic-cortical brain regions which project directly or indirectly to the striatum. In the present study, dopamine metabolism in subregions of the striatum was examined in rats with KA lesions after acute and chronic haloperidol or clozapine administration. The main findings was that the elevating effect of acute haloperidol treatment on the dopamine metabolite, DOPAC, was blocked in the nucleus accumbens shell and diminished in medial and laterodorsal caudate-putamen of the KA-lesioned rats. In addition, the elevating effects of both acute and chronic haloperidol treatment on dopamine turnover were attenuated in the laterodorsal caudate-putamen of KA-lesioned rats. The levels of dopamine, DOPAC, and HVA after chronic clozapine treatment were greater in KA-lesioned than control rats. These results indicate that dopaminergic responses to haloperidol may be diminished by limbic-cortical neuropathology, while such pathology does not significantly alter dopaminergic responses to clozapine.

Induction of Fos protein by antipsychotic drugs in rat brain following kainic acid-induced limbic-cortical neuronal loss.

Antipsychotic drugs increase expression of the immediate early gene, c-fos, in the striatum, nucleus accumbens and prefrontal cortex of rat brain. Since intracerebro-ventricular (i.c.v.) infusion of kainic acid (KA) produces loss of limbic-cortical neurons that project to these brain areas, we postulated that the c-fos responses to antipsychotics in these brain areas would be altered following i.c.v. KA administration. To produce limbic-cortical lesions, rats received i.c.v. infusions of either KA (4.5 nmol) or vehicle. Then, 25 28 days later, rats received 0.13, 0.35, or 1.5 mg/kg haloperidol, 6.3, 17.5, or 30.0 mg/kg clozapine, or saline. In both KA-lesioned and control animals, haloperidol produced greater increases in Fos protein immunoreactivity in the striatum than in limbic-cortical areas, while clozapine produced greater increases in Fos protein immunoreactivity in limbic-cortical areas than in the striatum. In both KA-lesioned and control animals, haloperidol and clozapine administration also produced similar dose-dependent increases in Fos protein immunoreactivity in the striatum and nucleus accumbens. However, the ability of clozapine to increase Fos protein immunoreactivity in the infralimbic prefrontal cortex was significantly enhanced in KA-lesioned rats compared to controls. Since limbic-cortical pathology has been implicated in the negative symptoms of schizophrenia, the enhanced effect of clozapine on limbic-cortical expression of c-fos in KA-lesioned rats may be relevant to understanding clozapine's unusual therapeutic actions in patients with schizophrenia.

The effects of kainic acid lesions on locomotor responses to haloperidol and clozapine.

Spontaneous and amphetamine-elicited locomotor activity in rats is reduced by most clinically effective antipsychotic drugs. We have recently demonstrated that intracerebroventricular infusion of kainic acid (KA), which produces cell loss in the hippocampus and other limbic-cortical brain regions, increases spontaneous and amphetamine-elicited locomotion. The present study determined if KA lesions alter the suppressive effects of the antipsychotic drugs, haloperidol and clozapine, on spontaneous and amphetamine-elicited locomotor behavior. Young adult male rats (70 days of age) received intracerebroventricular infusions of vehicle or KA, which produced hippocampal pyramidal cell loss in each rat and more variable cell loss or gliosis in the amygdala, piriform cortex, and laterodorsal thalamus. Thirty days post-surgery, lesioned and control rats were tested once a week for locomotor responses to drug treatments. As observed previously, spontaneous locomotor activity and hyperactivity elicited by amphetamine (1.50 mg/kg s.c.) were greater in lesioned animals than controls. In addition, the level of spontaneous activity and/or amphetamine-elicited hyperlocomotion observed in lesioned rats after haloperidol treatment (0.13, 0.35, or 1.50 mg/kg s.c.) was greater than that found in controls. Locomotor responses to low (6.30 mg/kg) and moderate doses of clozapine (20 mg/kg) were similar in lesioned and control rats, although lesioned rats were more active than controls following the administration of a high dose of clozapine (30 mg/kg). These data indicate that the hyperactivity associated with limbic-cortical lesions may be insensitive to reversal by haloperidol, yet uniquely sensitive to suppression by clozapine.

Kainic acid lesions enhance locomotor responses to novelty, saline, amphetamine, and MK-801.

Intracerebroventricular (i.c.v.) administration of kainic acid (KA) to rats produces neuronal loss in the hippocampus and other areas of the limbic system. The present study demonstrates that i.c.v. KA enhances the locomotor response to novelty and saline injection, as well as to amphetamine and MK-801. Sixteen to 18 days after i.c.v. administration of KA or vehicle, lesioned and control rats were placed in a novel cage, and locomotor activity and grooming were recorded for 30 min prior to and 60 min following a subcutaneous injection of saline, D-amphetamine, or MK-801. In response to the novel cage and after each injection, KA rats exhibited increased locomotor activity relative to controls. Grooming behavior was found to be elevated in the KA rats when compared to controls, but only in response to the novel cage and saline injection. The possibility that damage to the limbic system disrupts dopaminergic regulation of locomotor behavior is discussed, as well as implications for neuropathology in schizophrenia.

Kainic acid decreases hippocampal neuronal number and increases dopamine receptor binding in the nucleus accumbens: an animal model of schizophrenia.

Intracerebroventricular (i.c.v.) administration of kainic acid (KA) produces graded neuronal loss in the hippocampus and other regions of the medial temporal lobe. Many of these brain regions send excitatory projections to the nucleus accumbens, a dopaminergic brain area implicated in psychotomimetic and antipsychotic drug action. In the present study, neurochemical function in the nucleus accumbens and anterior caudate-putamen was examined one week after i.c.v. administration of 1.5, 4.5, or 6.6 nmol of KA. As expected, i.c.v. KA produced dose-dependent neuronal loss in the dorsal and ventral hippocampus. Extrahippocampal neuronal loss was also observed in the thalamus and piriform cortex in some of the KA-treated rats. While ambient levels of dopamine turnover and excitatory amino acids in the nucleus accumbens were unaltered by KA, administration of the highest KA dose elevated [3H]spiperone binding exclusively in the accumbens. Finally, behavioral hyperactivity was observed in KA-treated rats over a five-week period following i.c.v. administration. The pattern of neuronal loss, receptor upregulation, and behavioral hyperactivity found after i.c.v. KA administration may provide a useful animal model of the limbic neuropathology and neurochemical dysfunction associated with schizophrenia.

Effects of selective toxic lesions of cholinergic neurons of the laterodorsal tegmental nucleus on experimental seizures.

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.

Subcortical excitatory amino acid levels after acute and subchronic administration of typical and atypical neuroleptics.

The effects of three atypical neuroleptic compounds, clozapine, sulpiride, and (-)-3-(3-hydroxyphenyl)-N-n-propyl-piperidine ((-)-3-PPP) were compared to the effects of haloperidol and saline on excitatory amino acid levels in the rodent nucleus accumbens and corpus striatum after acute (1 day) and subchronic (28 days) treatment. Equivalent doses of each drug were determined by assessing their in vivo displacement of [3H]spiperone binding in the nucleus accumbens and corpus striatum. After acute treatment, all three atypical neuroleptics, but not haloperidol, produced a significant decrease in nucleus accumbens glutamate concentrations. Acute haloperidol treatment significantly elevated glutamate concentrations in the corpus striatum when compared to all three atypical drugs. After subchronic treatment, (-)-3-PPP significantly increased glutamate concentrations in the nucleus accumbens when compared to the effects of haloperidol and clozapine. There were no major between-group differences in glutamate levels after subchronic treatment in the corpus striatum. The effects of acute and subchronic neuroleptic administration on aspartate levels in the nucleus accumbens and corpus striatum were highly variable. These findings indicate that atypical and typical neuroleptics may alter subcortical excitatory amino acid levels in a site-specific manner.

Limbic-cortical neuronal damage and the pathophysiology of schizophrenia.

Neurobiological studies of patients with schizophrenia suggest that abnormalities of both anatomy and function occur in limbic-cortical structures. An anatomical circuit links the functioning of the ventral striatum (i.e., nucleus accumbens) with the hippocampus and other limbic-cortical structures where neurobiological abnormalities have been found. In animals, lesions of limbic-cortical neurons cause decreases in glutamatergic input to the nucleus accumbens and are also associated with decreases in presynaptic dopamine release, increases in the density of D2-like dopamine receptors, and insensitivity to the actions of dopamine antagonists such as haloperidol. These experiments suggest a plausible pathophysiology of schizophrenia, in that schizophrenic symptoms may be caused by an abnormal dopaminergic state brought about by a primary limbic-cortical lesion and deficits in glutamatergic inputs to the ventral striatum.

Maturation of locomotor and Fos responses to the NMDA antagonists, PCP and MK-801.

Antagonists at the N-methyl-D-aspartate (NMDA)-type glutamate receptor, such as phencyclidine (PCP) and dizocilpine (MK-801), are well-known to evoke increases in locomotor activity in adult rats and mice. However, little is known about the effects of NMDA antagonists on locomotor activity as a function of development. The present study examined locomotor responses to PCP or MK-801 in male rats of varying ages and found that prepubertal rats were more sensitive to the locomotor-elevating effects of PCP (1.5 mg/kg and 3. 0 mg/kg, s.c.) than were adults. Locomotor responses to MK-801 (0.1 and 0.2 mg/kg, s.c.) were not dependent on age. The age-dependent response to PCP may be related to developmental events in the motor cortex, since more Fos-immunoreactive neurons were observed in the motor cortex of prepubertal animals after PCP administration relative to adult animals. An opposite pattern of age-dependent Fos responses was observed in the posterior retrosplenial cortex. The results suggest that locomotor responses to NMDA antagonists can be influenced in an age- and drug-dependent manner and that maturational events in the motor cortex may modify responses to PCP.