Effects of apolipoprotein E, beta-very low density lipoproteins, and cholesterol on the extension of neurites by rabbit dorsal root ganglion neurons in vitro.

Previous studies suggest that during nerve regeneration apoE acts as a lipid transport protein that assists in the rapid initial extension of axons and then in their myelination. To determine whether apoE and/or apoE-containing lipoproteins can modulate axon growth, we assessed their effect on the out-growth of neurites from neurons in mixed cultures of fetal rabbit dorsal root ganglion cells in vitro. Incubation with beta-very low density lipoprotein (beta-VLDL) particles, which are rich in apoE and cholesterol, increased neurite outgrowth and branching. Unesterified cholesterol added to the cultures had a similar, but less pronounced, effect. These data suggest that cholesterol might be the component responsible for the enhanced neurite growth. In contrast, purified, lipid-free apoE added to the cultures reduced neurite branching. Neurite branching was also reduced when purified apoE was added along with beta-VLDL or cholesterol; however, the striking finding was that under these conditions the neurites extended farther from the neuronal cell body. Dorsal root ganglion cells were examined for the presence of receptors for native and apoE-enriched beta-VLDL. Immunocytochemistry, ligand blots, 45Ca2+ blots, and studies of the interaction of the cells with fluorescent lipoproteins provided evidence of two types of receptors for apoE-containing lipoproteins on neurons: the low density lipoprotein (LDL) receptor, which binds native beta-VLDL, and the LDL receptor-related protein, which binds apoE-enriched beta-VLDL. These findings indicate that apoE may play two complementary roles in neurite outgrowth. When complexed with lipoproteins, apoE stimulates neurite growth by the receptor-mediated delivery of cholesterol and perhaps other components necessary for neurite outgrowth. When apoE as a free protein is added together with apoE-containing lipoproteins, apoE decreases neurite branching and promotes neurite extension away from the cell body. These actions, which would be complementary in promoting target-directed nerve growth in vivo, provide the first direct evidence that apoE and apoE-containing lipoproteins can modulate the outgrowth of neuronal processes.

Milacemide, a glycine prodrug, enhances performance of learning tasks in normal and amnestic rodents.

The N-methyl-D-aspartate receptor complex appears to play an important role in processes of learning and memory. The presence of a glycine modulatory site at this complex has recently been established and suggests that glycinergic neurotransmission may influence these cognitive functions. Increasing glycine concentrations in the brain by administration of a glycine prodrug, milacemide, is shown here to enhance performance of a shock-motivated passive avoidance task in rats, and to reverse drug-induced amnesia in a spontaneous alternation paradigm in mice. Prevention of the metabolism of milacemide to glycine by pretreatment with MAO-B inhibitors not only prevents the memory-enhancing effects of the drug, but appears to have a deleterious effect on memory formation suggesting an action of the prodrug itself on the brain. These studies indicate a role of glycinergic neurotransmission in memory processes, and support the therapeutic potential of glycinergic drugs in memory impairment.

D-cycloserine, a positive modulator of the N-methyl-D-aspartate receptor, enhances performance of learning tasks in rats.

Glycine has recently been shown to positively modulate the N-methyl-D-aspartate (NMDA) subclass of acidic amino acid receptors which are important in neural pathways involved in learning and memory. We report that d-cycloserine (DCS), an antimycobacterial agent known to cross the blood-brain barrier, binds with high affinity to this glycine modulatory site, functions as a positive modulator, and facilitates performance of learning tasks in rats. In addition, DCS appears to be a potent cognitive enhancer at doses lower than those required for antibacterial activity. Based on these data, we propose that modulation of NMDA receptors via glycinergic mechanisms may be a means of influencing cognitive processes.

Selective memory impairment by phencyclidine in rats.

Phencyclidine (PCP) users sometimes report lack of recall of events occurring while they are under the influence of the drug. The present experiment was designed to test whether rats remember information learned after PCP administration. Rats were trained to choose one arm of a T-maze to obtain a food reward. The following day they were injected with either PCP (1 mg/kg) or vehicle and trained to choose the opposite arm for a reward (reversal learning). A third group of rats received neither injections nor training on the second day. On the third day, all rats were tested for their preference of maze arms. Rats who had been injected with saline before reversal learning chose the arm rewarded during the reversal, while rats receiving PCP on the second day chose randomly. The rats which did not learn the reversal chose the arm learned on day 1. These results indicate that while PCP did not interfere with the rats ability to learn, it interfered with long-term storage of information.

A developmental influence of substance P on its receptors.

Events which alter the chemical environment of the developing nervous system have long-term consequences for neural function and behavior. Neonatal exposure to a peptide neurotransmitter, substance P (SP), increased the number of SP binding sites in the adult rat salivary glands, without altering their affinity. The neonatal treatment also increased the binding capacity for SP of several brain regions, but had no effect on SP levels in the brain. The increase in SP binding sites may be responsible for increases in sensitivity to SP in adults after neonatal treatment with the peptide.

Phencyclidine. Physiological actions, interactions with excitatory amino acids and endogenous ligands.

Phenycyclidine (PCP) produces many profound effects in the central nervous system. PCP has numerous behavioral and neurochemical effects such as inhibiting the uptake and facilitating the release of dopamine, serotonin, and norepinephrine. PCP also interacts with sigma, mu opioid, muscarinic, and nicotinic receptors. However, the psychotomimetic effects induced by PCP are believed to be mediated by specific PCP receptors, where PCP binds with greater potency than sigma compounds. Electrophysiological, behavioral, and neuro-chemical evidence strongly suggests that at least some of the many PCP actions result from antagonism of excitatory amino acid-induced responses via PCP receptors. The recent isolation and partial characterization of the alpha and beta endopsychosins and the identification of other endogenous ligands for the PCP and sigma receptors, is another promising area of research in the elucidation of the physiological role of an endogenous PCP and sigma system.

Secretin receptors in the rat kidney: adenylate cyclase activation and renal effects.

In previous studies it has been demonstrated that pharmacological administration of secretin can alter urine output. Whether the effect is due to a direct action on kidney was investigated by examining the effect of secretin on renal output, and determining whether there were secretin receptors and a secretin sensitive adenylate cyclase in the kidney. Secretin had an antidiuretic action on kidney when administered intravenously to anesthetized hydrated rats. In addition, binding sites for (125I)-secretin, and a secretin sensitive adenylate cyclase were identified in rat kidney. Binding was saturable and reversable and was half maximally inhibited by 1 X 10(-7) M synthetic porcine secretin. Autoradiographic studies revealed a high density of secretin binding sites in the outer medulla of the kidney, a region that is composed mainly of the thick ascending limb of the loop of Henle, and is also the major site of action for the antidiuretic hormone, vasopressin. The data indicate that a functional secretin receptor system exists in kidney which may have a physiological role in regulating urine output.

Neuropeptide effects on brain development.

Peptides which act as neurotransmitters or neuromodulators in the adult nervous system exert influences on brain development. Altering the levels of neuropeptide to which a developing animal is exposed produces a wide variety of physiological and behavioral effects which are apparent in adulthood. Many of the developmental effects involve neural systems in which the neuropeptide plays a role in the adult.

Modulation of brain development by morphine: effects on central motor systems and behavior.

Morphine administration to neonatal rats on days 1-7 after birth produced long-term changes in behavior and brain function. The pups were smaller than saline-treated littermates and showed retarded motor development. As adults, the morphine-treated rats had impaired motor coordination, altered gait, and altered patterns of activity in an open field. Several brain regions of the adult rats, including motor areas, had decreased metabolic activity as measured by the 2-deoxy-glucose technique, suggesting decreased functional activity in these areas. These results may be relevant to findings that children exposed in utero to narcotics tend to have impaired motor development.

Neonatal exposure to vasopressin decreases vasopressin binding sites in the adult kidney.

Previous experiments showed that rats injected with vasopressin (AVP) during the first seven days after birth were less sensitive as adults to the antidiuretic effects of AVP than were control rats. In the present experiment, binding sites for AVP were measured in the kidneys of similarly treated adult rats. Neonatal exposure to AVP significantly decreased the number of binding sites in the adults, but did not affect the binding affinity of the sites. It is concluded that neonatal exposure to AVP which produces a long-lasting decrease in responsiveness of the kidney to AVP is correlated with a reduction in the number of AVP binding sites in the tissue.

Substance P administration to neonatal rats increases adult sensitivity to substance P.

Substance P (SP) administered subcutaneously to neonatal rats on days 1-7 after birth produced long-term physiological changes. The changes included altered pain perception and increased sialogogic response to SP, although the hypotensive response to SP was unchanged. Early exposure to the peptide therefore influenced development, particularly with respect to two systems in which SP is physiologically active. Both changes may reflect an increased sensitivity of these systems to the effects of SP.

Vasopressin administration to neonatal rats reduces antidiuretic response in adult kidneys.

Neonatal rats who had been given injections of vasopressin on days 1-7 after birth exhibited polyuria as adults. In vivo antidiuresis bioassays demonstrated that their kidneys were deficient in their ability to concentrate urine in response to stimulation with vasopressin. The kidneys also showed a reduction in vasopressin-induced cyclic AMP production, although parathyroid hormone- and calcitonin-induced levels were normal. This suggests a specific deficit in vasopressin receptor-adenylate cyclase function. In contrast, the neonatal treatment had no effect on the sensitivity of the adult vasculature to the hypertensive effects of vasopressin. These results show that short exposures to high levels of vasopressin early in development can produce a long-term defect in vasopressin responsiveness that is specific to the kidney.

Secretin modulation of behavioral and physiological functions in the rat.

The effect of secretin on behavioral and physiological functions in the rat was investigated. Secretin injected intracerebroventricularly (ICV) significantly increased defecation and decreased novel-object approaches in rats. The peptide showed no significant effects on stereotypic behavior (gnawing, grooming and rearing), open-field locomotor activity however was significantly decreased, an effect that was probably due to a decreased propensity for the rats to initiate locomotor responses. In addition, secretin showed significant effects on respiration rate in anesthetized rats. When the peptide was injected in the lateral ventricle a decrease in respiration rate occurred, but when the brain was perfused from the lateral ventricle to the cisterna magna increases in respiration rate occurred. These data, combined with the facts that secretin and secretin receptors have been identified in the brain indicate that secretin may play a neurotransmitter or neuroregulator role in the central nervous system.

Behavioral effects of [4-norleucine, 7-D-phenylalanine]-alpha-melanocyte-stimulating hormone.

The behavioral effects of alpha-melanocyte stimulating hormone (alpha-MSH) were compared to an alpha-MSH analogue that had a norleucine substituted for methionine in the four position and a D-phenylalanine substituted for L-phenylalanine in the seven position. [Nle4, D-Phe7]-alpha-MSH has previously been shown to be a superpotent agonist on melanocytes [17]. The present experiments indicate that [Nle4, D-Phe7]-alpha-MSH is equipotent to alpha-MSH in inducing grooming when administered intraventricularly. In contrast, the analogue has the opposite effect of alpha-MSH on performance of a visual discrimination task. alpha-MSH improves visual performance whereas [Nle4, D-Phe7]-alpha-MSH attenuates such performance. The contrasting activities of [Nle4, D-Phe7]-alpha-MSH on the physiological processes described suggest that this analogue may interact with three distinct melanotropin receptors in different ways. On melanocyte receptors the melanotropin analogue is a superagonist, on CNS melanotropin receptors involved in grooming it is equipotent to alpha-MSH, and on CNS receptors involved in attention, learning and memory [Nle4, D-Phe7]-alpha-MSH may be an antagonist of endogenous melanotropin.

Extra-hippocampal projections of CCK neurons of the hippocampus and subiculum.

A population of neurons in the hippocampus and subiculum contains cholecystokinin (CCK). Following transection of the dorsal fornix, a major afferent pathway of the hippocampus and associated structures. CCK levels were reduced in the septum and hypothalamus. A microdissection analysis indicated that the loss of CCK occurred in nuclei receiving direct projections from the hippocampus and subiculum, suggesting that CCK-containing neurons in the hippocampus and subiculum project to extrahippocampal regions.

Effects of time and experience on hippocampal neurochemistry after damage to the CA3 subfield.

Bilateral injections of kainic acid into the hippocampal CA3 subfield destroyed the CA 3 pyramidal cells and produced a behavioral impairment, an inability to solve spatial maze problems. The behavior recovered, however, with daily experience in a maze task, and the rate of recovery was accelerated by additional daily experience. This recovery of function could be the result of compensatory changes in the distribution or function of the various hippocampal pathways. In the present experiment, this possibility was investigated neurochemically. Five putative neurotransmitters or their synthetic enzymes were measured in dissected regions of the hippocampal formation. Both the long-term effects of the lesions and the effects of behavioral training were determined. A number of alterations in hippocampal neurochemical systems were detected. Acute changes due to the lesions included a widespread loss of glutamate, and regionally specific decreases in glutamic acid decarboxylase (GAD) activity and cholecystokinin (CCK) and norepinephrine (NE) concentrations. Long-term changes included a decline in choline acetyltransferase (ChAT) activity throughout the hippocampal formation, and increases in NE in certain regions. Behavioral testing prevented the decline of ChAT activity, and increased the concentrations of GAD and CCK. The neurochemical conditions present at the time when trained rats recovered behavioral function may indicate the crucial conditions for the occurrence of the behavior.

Choline acetyltransferase activity in the nucleus tractus solitarius: regulation by the afferent vagus nerve.

The influence of nodose ganglionectomy or transection of the peripheral branches of the afferent vagus nerve on choline acetyltransferase (ChAT) activity in the nucleus tractus solitarius (NTS) was studied. ChAT activity was reduced in the microdissected caudal and intermediate portions of the NTS in vagotomized as well as ganglionectomized rats. However, only the ganglionectomy resulted in the degeneration of medullary nerve fibers. These results suggest that the changes in ChAT activity in the NTS are independent of neuronal degeneration and may be due to transynaptic modulation of ChAT activity by afferent vagal impulses. The presence of ChAT in the sensory nodose projection to the NTS, however, cannot be ruled out.

Alpha-melanocyte stimulating hormone facilitates learning of visual but not auditory discriminations.

The neuropeptide alpha-MSH has been proposed to influence learning and memory by increasing visual attention. To test the possibility that MSH selectively affects visual learning, rats were tested in learning tasks in which the cues were either visual or auditory. Maze and bar-press tasks were used. MSH administration increased the rate of learning of the visual tasks, regardless of the task difficulty or the type of response required of the rat. MSH had no effect on the rate of learning of the auditory tasks. These results support the hypothesis that MSH facilitates learning by influencing some aspect of visual information processing.