Prenatal Protein Malnutrition Leads to Hemispheric Differences in the Extracellular Concentrations of Norepinephrine, Dopamine and Serotonin in the Medial Prefrontal Cortex of Adult Rats.

Exposure to prenatal protein malnutrition (PPM) leads to a reprogramming of the brain, altering executive functions involving the prefrontal cortex (PFC). In this study we used in vivo microdialysis to assess the effects of PPM on extracellular concentrations of norepinephrine (NE), dopamine (DA) and serotonin (5-HT) bilaterally in the ventral portion of the medial prefrontal cortex (vmPFC; ventral prelimbic and infralimbic cortices) of adult Long-Evans rats. Female Long-Evans rats were fed either a low protein (6%) or adequate protein diet (25%) prior to mating and throughout pregnancy. At birth, all litters were culled and fostered to dams fed a 25% (adequate) protein diet. At 120 days of age, 2 mm microdialysis probes were placed into left and right vmPFC. Basal extracellular concentrations of NE, DA, and 5-HT were determined over a 1-h period using HPLC. In rats exposed to PPM there was a decrease in extracellular concentrations of NE and DA in the right vmPFC and an increase in the extracellular concentration of 5-HT in the left vmPFC compared to controls (prenatally malnourished: N = 10, well-nourished: N = 20). Assessment of the cerebral laterality of extracellular neurotransmitters in the vmPFC showed that prenatally malnourished animals had a significant shift in laterality from the right to the left hemisphere for NE and DA but not for serotonin. In a related study, these animals showed cognitive inflexibility in an attentional task. In animals in the current study, NE levels in the right vmPFC of well-nourished animals correlated positively with performance in an attention task, while 5-HT in the left vmPFC of well-nourished rats correlated negatively with performance. These data, in addition to previously published studies, suggest a long-term reprogramming of the vmPFC in rats exposed to PPM which may contribute to attention deficits observed in adult animals exposed to PPM.

Prenatal malnutrition leads to deficits in attentional set shifting and decreases metabolic activity in prefrontal subregions that control executive function.

Globally, over 25% of all children under the age of 5 years experience malnutrition leading to cognitive and emotional impairments that can persist into adulthood and beyond. We use a rodent model to determine the impact of prenatal protein malnutrition on executive functions in an attentional set-shifting task and metabolic activity in prefrontal cortex (PFC) subregions critical to these behaviors. Long-Evans dams were provided with a low (6% casein) or adequate (25% casein) protein diet 5 weeks before mating and during pregnancy. At birth, the litters were culled to 8 pups and fostered to control dams on the 25% casein diet. At postnatal day 90, prenatally malnourished rats were less able to shift attentional set and reverse reward contingencies than controls, demonstrating cognitive rigidity. Naive same-sexed littermates were assessed for regional brain activity using the metabolic marker (14)C-2-deoxyglucose (2DG). The prenatally malnourished rats had lower metabolic activity than controls in prelimbic, infralimbic, anterior cingulate, and orbitofrontal cortices, but had comparable activity in the nearby piriform cortex and superior colliculus. This study demonstrates that prenatal protein malnutrition in a well-described animal model produces cognitive deficits in tests of attentional set shifting and reversal learning, similar to findings of cognitive inflexibility reported in humans exposed to early childhood malnutrition.

A microdialysis study of the medial prefrontal cortex of adolescent and adult rats.

The medial prefrontal cortex (mPFC) of the rat has become a key focus of studies designed to elucidate the basis of behavior involving attention and decision-making, i.e. executive functions. The adolescent mPFC is of particular interest given the role of the mPFC in impulsivity and attention, and disorders such as attentional deficit disorder. In the present study we have examined the basal extracellular concentrations of the neurotransmitters 5-hydroxytryptamine (5-HT), dopamine (DA) and norepinephrine (NE) in the ventral portion of the mPFC (vmPFC) in both adolescent (post-natal day 45-50) and adult, and male and female rats using in vivo microdialysis. We have also examined both the left and right vmPFCs given reports of laterality in function between the hemispheres. Basal extracellular concentrations of 5-HT differed significantly between male and female rats. Extracellular DA also differed significantly between male and female rats and between the left and the right vmPFC in adult males. No differences were seen in basal extracellular NE. There was a significant age difference between groups in the laterality of extracellular NE levels between right and left vmPFC. Infusion of 100 µM methamphetamine through the dialysis probe increased the extracellular concentration of all the monoamines although there were no differences between groups in methamphetamine stimulated release. The findings from this study demonstrate that there are differences in monoaminergic input to the mPFC of the rat based on age, gender and hemisphere. This work sets the neurochemical baseline for further investigations of the prefrontal cortex during development.

Functional interrelations between nucleus raphé dorsalis and nucleus raphé medianus: a dual probe microdialysis study of glutamate-stimulated serotonin release.

Dual-probe in vivo microdialysis was used to explore the relationships between the two midbrain raphé nuclei, raphé dorsalis (DRN) and raphé medianus (MRN). Infusion of the excitatory neurotransmitter glutamate (10 mM) into the dorsal raphé nucleus produced a large increase in the extracellular 5-HT (5-HT(ext)) in the dorsal raphé (1400% of control values) that was limited to the time of infusion. This was followed by a significant decrease in extracellular 5-HT below baseline levels that continued for the duration of the experiment (3 h). Extracellular 5-HT (5-HT(ext)) was also increased to 500% of control values in the median raphé nucleus following infusion of 10 mM glutamate (GLU) into the dorsal raphé nucleus. Infusion of the competitive NMDA receptor antagonist AP5 prior to and during infusion of GLU into the DRN resulted in a decrease in the response to GLU in the DRN and an antagonism of the increase of 5-HT(ext) in the MRN. Infusion of 10mM GLU into the lateral midbrain tegmentum, an area of the brain just lateral to the DRN, also increased 5-HT(ext) in the probe in the lateral midbrain tegmentum (900% of control) but did not alter 5-HT(ext) in the MRN. When glutamate was infused into the MRN, 5-HT(ext) was also increased to 1400% of control in a time course similar to that seen with infusion of GLU into the DRN. Infusion of glutamate into the MRN, however, did not alter the 5-HT(ext) in the DRN. These data suggest a serotonergic innervation of the median raphé nucleus by the dorsal raphé nucleus. A reciprocal innervation from the median raphé to the dorsal raphé is not mediated by glutamate, does not appear to be serotonergic, and does not regulate extracellular serotonin in the dorsal raphé.

Stress-induced changes in extracellular dopamine and serotonin in the medial prefrontal cortex and dorsal hippocampus of prenatally malnourished rats.

Prenatal protein malnutrition continues to be a significant problem in the world today. Exposure to prenatal protein malnutrition increases the risk of a number of neuropsychiatric disorders in adulthood including depression, schizophrenia and attentional deficit disorder. In the present experiment, we have examined the effects of stress on extracellular serotonin (5-HT) and dopamine in the medial prefrontal cortex and dorsal hippocampus of rats exposed in utero to protein malnutrition. The medial prefrontal cortex and dorsal hippocampus were chosen as two limbic forebrain regions involved in learning and memory, attention and the stress response. Extracellular 5-HT and dopamine were determined in the medial prefrontal cortex and dorsal hippocampus of adult male Sprague-Dawley rats using dual probe in vivo microdialysis. Basal extracellular 5-HT did not differ between malnourished and well-nourished controls in either the medial prefrontal cortex or the dorsal hippocampus. Basal extracellular dopamine was significantly decreased in the medial prefrontal cortex of malnourished animals. Restraint stress (20 m) produced a significant rise in extracellular dopamine in the medial prefrontal cortex of well-nourished rats but did not alter release in malnourished rats. In malnourished rats, stress produced an increase in 5-HT in the hippocampus, whereas stress produced a decrease in 5-HT in the hippocampus of well-nourished rats. These data demonstrate that prenatal protein malnutrition alters dopaminergic neurotransmission in the medial prefrontal cortex as well as alters the dopaminergic and serotonergic response to stress. These changes may provide part of the bases for alterations in malnourished animals' response to stress.

The limbic brain: continuing resolution.

This paper presents an overview of the limbic brain and its distributed sub-systems. The extent of the limbic system has expanded in recent years. Among the brain areas that we now argue should be included in the extended limbic system are the medial prefrontal cortex, the insular cortex as well as the lower brainstem and spinal cord. In addition the limbic forebrain and limbic midbrain may be divided into medial and lateral divisions both anatomically and physiologically. This serves as an introduction to the papers that follow.

A review of systems and networks of the limbic forebrain/limbic midbrain.

Evolutionarily older brain systems, such as the limbic system, appear to serve fundamental aspects of emotional processing and provide relevant and motivational information for phylogenetically more recent brain systems to regulate complex behaviors. Overall, overt behavior is, in part, determined by the interactions of multiple learning and memory systems, some seemingly complementary and some actually competitive. An understanding of limbic system function in emotion and motivation requires that these subsystems be recognized and characterized as extended components of a distributed limbic network. Behavioral neuroscientists face the challenge of teasing apart the contributions of multiple overlapping neuronal systems in order to begin to elucidate the neural mechanisms of the limbic system and their contributions to behavior. One major consideration is to bring together conceptually the functions of individual components of the limbic forebrain and the related limbic midbrain systems. For example, in the rat the heterogeneous regions of the prefrontal cortex (e.g., prelimbic, anterior cingulate, subgenual cortices and orbito-frontal areas) make distinct contributions to emotional and motivational influences on behavior and each needs consideration in its own right. Major interacting structures of the limbic system include the prefrontal cortex, cingulate cortex, amygdaloid nuclear complex, limbic thalamus, hippocampal formation, nucleus accumbens (limbic striatum), anterior hypothalamus, ventral tegmental area and midbrain raphe nuclei; the latter comprising largely serotonergic components of the limbic midbrain system projecting to the forebrain. The posterior limbic midbrain complex comprising the stria medullaris, central gray and dorsal and ventral nuclei of Gudden are also key elements in the limbic midbrain. Some of these formations will be discussed in terms of the neurochemical connectivity between them. We put forward a systems approach in order to build a network model of the limbic forebrain/limbic midbrain system, and the interactions of its major components. In this regard, it is important to keep in mind that the limbic system is both an anatomical entity as well as a physiological concept. We have considered this issue in detail in the introduction to this review. The components of these systems have usually been considered as functional units or 'centers' rather than being components of a larger, interacting, and distributed functional system. In that context, we are oriented toward considerations of distributed neural systems themselves as functional entities in the brain.

Modulation of 5-HT release in the hippocampus of 30-day-old rats exposed in utero to protein malnutrition.

Previous in vivo microdialysis studies have shown increased spontaneous release of 5-HT in the hippocampus of adult behaving rats exposed to prenatal protein malnutrition. Furthermore, behavioral studies have shown that adolescent rats (PD30) that have been prenatally protein malnourished demonstrate an increased sensitivity to the benzodiazepine chlordiazepoxide (CDP). Given this altered sensitivity to benzodiazepines in adolescent malnourished rats, the present study was designed to test the hypothesis that the increased release of 5-HT in the hippocampus is present in adolescent rats and that this release is modulated by CDP. An altered release of 5-HT at PD30 would suggest an early developmental change associated with prenatal malnutrition. PD30 rats were implanted with microdialysis probes into the dorsal hippocampus and 5-HT release was monitored before and after administration of CDP. As previously reported in adult rats, release of 5-HT was significantly elevated in the dorsal hippocampus of PD30 rats as compared to well-nourished 30-day-old controls. Administration of CDP did not affect the release of 5-HT from the hippocampal formation of well-nourished rats but significantly decreased the elevated release of 5-HT in the malnourished rats. Following CDP, 5-HT release in the malnourished rats was at the same levels as release in well-nourished animals. Benzodiazepines have been reported to decrease extracellular 5-HT in stressed rats but not in unstressed rats. Thus, the elevated 5-HT release in the hippocampus in rats exposed to prenatal protein malnutrition may be associated with an increased response to stress. These data support other data that prenatal protein malnutrition alters the response to stressful stimuli possibly through changes in the GABAergic and/or serotonergic systems.

Effects of prenatal protein malnutrition on the hippocampal formation.

In this review we have assessed the effects of prenatal protein malnutrition on the hippocampal formation of the developing brain. In investigating this insult in the hippocampal neuronal model we have concentrated on aspects of enhanced inhibition we have shown in our earlier studies. Since this involves particular attention to the GABAergic interneurons we have examined the complex interneuronal networks of the hippocampal formation and their neurotransmitter afferent inputs, particularly the serotonergic system from the midbrain raphé nuclei. A variety of combinations of specialized interneurons are discussed in terms of how malnutrition insults perturb function in these inhibitory and disinhibitory networks. Pathological enhancement of inhibition manifests itself by diminished plasticity, alterations in theta activity and deficits in long-term learning behaviors. Long-term inhibition in select GABA interneuron systems may form a major derangement seen following prenatal protein malnutrition. The focus of this study is to relate enhanced inhibition to the several forms of inhibitory systems present in the hippocampal formation and develop hypotheses as to the primary derangements that may account for pathological inhibition in prenatal malnutrition.