John P. Bruno

Apparent sprouting of striatal serotonergic terminals after dopamine-depleting brain lesions in neonatal rats

Near-total dopamine-depleting brain lesions produced in 3-day-old rats by intracerebroventricular injection of the neurotoxin 6-hydroxydopamine led to pronounced increases in striatal serotonin (5-HT) and 5-hydroxyindoleacetic acid contents 1-8 months later. This effect was associated with an increase in in vitro high affinity 5-HT uptake, suggesting that proliferation of new serotonergic terminals had occurred within the striatum. No such effect was obtained when comparable brain lesions were produced in adult rats.

Rats given dopamine-depleting brain lesions as neonates are subsensitive to dopaminergic antagonists as adults

Extensive damage to central dopamine (DA)-containing neurons are known to produce akinesia and sensory neglect when the lesions are made in adult rats. Similar behavioral impairments occur when dopaminergic function is disrupted temporarily by DA receptor blocking agents, and brain-damaged rats are particularly sensitive to the effects of those drugs. The present experiments offer a striking contrast to these well-accepted findings that central DA-containing neurons are critical to the initiation of voluntary movement. After near-total destruction of the dopaminergic neurons in 3-day-old rats, there were no conspicuous behavioral dysfunctions at any time during the subsequent 5-8 months, even when the animals were given large doses of DA receptor blocking agents. These findings suggest that some other neuronal system had replaced the absent dopaminergic neurons in the central control of movement.

Effect of dopamine-depleting brain lesions in rat pups: role of striatal serotonergic neurons in behavior

Previous results from our laboratory have demonstrated that 3-day-old rats given dopamine (DA)-depleting brain lesions are spared the severe behavioral dysfunctions seen after comparable brain damage in adults. This behavioral sparing is accompanied by a sprouting of serotonin (5-HT)-containing neurons in the striatum. The present results extend these observations by demonstrating that rats given the brain lesions as 15- or 27-day-olds continue to suckle, wean, and grow into adulthood without exhibiting any obvious behavioral dysfunctions, yet striatal 5-HT levels do not increase. Moreover, combined destruction of DA- and 5-HT-containing neurons in 3-day-old rat pups also produced no obvious behavioral dysfunctions. These and other results indicate that increases in striatal 5-HT are not necessary for the behavioral sparing observed after DA-depleting brain lesions in neonatal rats.

Effect of dopamine-depleting brain lesions on suckling and weaning in rats.

Rats given large dopamine-depleting brain lesions as adults exhibit severe impairments in ingestive behavior and sensorimotor function. In contrast to these well-known effects, virtually complete destruction of central dopaminergic neurons produced no such dysfunctions when it occurred in neonates. Indeed, rats continued to suckle and grow, albeit somewhat more slowly, and they could be weaned readily when they were 27 days old. Although most brain-damaged animals did not survive weaning when they were 18 days old, whereas controls exhibited no difficulty, this failure appears to be the consequence of their reduced body weight and related inability to maintain body temperature in a relatively cool environment (22 degrees C). Such premature weaning occurred more successfully when growth was stimulated by rearing brain-damaged pups in small litters or when ambient temperatures were raised to 31 degrees C so as to minimize heat loss. These results demonstrate that the effects of near-total dopamine-depleting brain lesions are considerably less severe when they occur in infants than when they occur in adults, and, consequently, they reveal a capacity for neural plasticity during development that is no longer present at maturity.

Inhibition of striatal acetylcholine release by serotonin and dopamine after the intracerebral administration of 6-hydroxydopamine to neonatal rats

The intraventricular administration of 6-hydroxydopamine (6-OHDA) depletes the striatum of dopamine (DA). When given to rat pups at an early age, the toxin also increases striatal serotonin (5-HT) content. In the accompanying report we observed that endogenous 5-HT, like DA, exerts an inhibitory influence on the release of acetylcholine (ACh) from striatal slices prepared from control animals and that the extent of this inhibition is related to the degree of serotonergic innervation of the region being examined. To determine whether this hyperinnervation was accompanied by an increase in serotonergic influence on ACh release, striatal slices were prepared from adult rats, preincubated with [3H]choline, superfused, and exposed to electrical field stimulation. The efflux of tritium into the superfusate was used as a measure of ACh release. In confirmation of previous reports, we observed that direct and indirect agonists of DA and 5-HT both reduced ACh overflow from control slices, whereas overflow was increased by antagonists of these amines. Slices prepared from rats given 6-OHDA-induced lesions as adults were responsive to each of these pharmacological manipulations, as well. In contrast, ACh overflow from slices prepared from animals lesioned with 6-OHDA as neonates was not modified by either dopaminergic or serotonergic drugs. These results suggest that the serotonergic hyperinnervation of striatum produced by neonatal 6-OHDA is accompanied by a loss of the inhibitory influence of endogenous 5-HT and DA on striatal ACh release and, thus, provide no evidence for a role for either transmitter in the behavioral sparing associated with such lesions.

Inhibition of striatal acetylcholine release by endogenous serotonin

We have examined the hypothesis that endogenous serotonin (5-HT) exerts an inhibitory influence on the release of acetylcholine (ACh) in striatum. Striatal slices were prepared from adult rats, preincubated with [3H]choline, superfused, and exposed to electrical field stimulation. The stimulation-induced overflow of tritium into the superfusate was used as a measure of ACh release. We observed that fluoxetine, an inhibitor of 5-HT uptake, reduced ACh overflow in slices prepared from caudal striatum, an area of high 5-HT concentration, but not in slices from rostral striatum, an area of low 5-HT concentration. Moreover, methysergide, a 5-HT antagonist, increased ACh efflux in caudal but not rostral striatum. Finally, direct activation of 5-HT receptors with the 5-HT agonist, quipazine, inhibited stimulation-induced ACh overflow in both rostral and caudal striatum. These results suggest that endogenous 5-HT normally is capable of inhibiting striatal ACh release, and that the extent of the modulation is related to the degree of serotonergic innervation. In addition, 5-HT receptors capable of modulating ACh release are present in 5-HT-poor rostral striatum, as well as in 5-HT-rich caudal striatum.

Rats given dopamine-depleting brain lesions as neonates do not respond to acute homeostatic imbalances as adults.

Rats given near-total dopamine-depleting brain lesions as neonates exhibit none of the severe sensorimotor or ingestive dysfunctions that result when adult rats are given comparable lesions. These experiments showed that the apparent sparing of function is incomplete; when tested as adults, the brain-damaged animals did not eat in response to acute glucoprivation, nor did they drink water and saline in response to hypovolemia. Indeed, after receiving these homeostatic challenges, the animals became much less able to simply move about. In contrast to these marked effects, the dopamine-depleted rats ate normally after glucoprivation when they were handled hourly instead of being left alone. Similarly, administration of caffeine permitted them to drink while hypovolemic. These and other observations suggest that some central neurons other than those containing dopamine are responsible for maintaining behavioral activation in rats after neonatal destruction of dopaminergic fiber systems in the brain and that the homeostatic challenges compromise their function and thereby disrupt behavior.

Neonatal dopamine depletions spare lateral hypothalamic stimulation reward in adult rats

Previous research has shown that adult rats sustaining near-total depletions of striatal dopamine (DA) as neonates exhibit few of the profound deficits in ingestion and sensory-motor behavior seen in comparably lesioned adults. This study extends these findings to another realm of DA-related behavior, reward function. In a rate-frequency curve-shift measurement paradigm, reward effectiveness of lateral hypothalamic brain stimulation was shown to be normal in adult rats depleted of brain DA as neonates. However, impairments were seen in rapid-initiation operant performance. Neonatally DA-depleted rats were also shown to be subsensitive to the DA receptor antagonist pimozide, suggesting that activity within undamaged DA neurons is not necessary for the elicitation of hypothalamic self-stimulation reward.

Sprouting of striatal neurons following neonatal 6-hydroxydopamine

A genomic l i b ra r y of s ingle copy rat DNA was prepared in phage Ml3. Total rat DNA was digested with the res t r i c t i on endonuclease Mbol. Unique Sequence DNA was isolated and l igated in to the Bam HI s i te of the rep l i ca t i ve form of the bacteriophage Ml3 mp7. Single stranded DNA from ind iv idua l recombinant Ml3 mp7 clones was digested with Sau 3A, which p re fe ren t i a l l y removes the rat DNA insert , label led with 32p by polynucleotide kinase, and hybridized to adult brain to ta l RNA. Clones containing sequences homologous to brain t ranscr ip ts were determined by a var ia t ion of the t ranscr ip t ion mapping technique. About 20% of these s ingle stranded clones were homologous to adult rat brain RNA, which agrees with our ea r l i e r RNA complexity data (Chikaraishi et a l , Cell 13, 1978). Also as expected, these clones correspond to rare brain t ranscr ip ts (O.05-1~opy per c e l l ) . These sequences were fu r ther characterized in terms of t ranscr ip t ion in heterologous tissues and in new born brain. At least two of the clones are spec i f i ca l l y transcribed in brain. The t ranscr ip t ion patterns of these cloned DNA fragments may reveal developmental patterns of gene expression.