Chronic stress exposure has been reported to induce dendritic remodeling in several brain regions, but it is not known whether individual neural circuits show distinct patterns of remodeling. The current study tested the hypothesis that the projections from the infralimbic (IL) area of the medial prefrontal cortex (mPFC) to the basolateral nucleus of the amygdala (BLA), a pathway relevant to stress-related mental illnesses like depression and post-traumatic stress disorder, would have a unique pattern of remodeling in response to chronic stress. The retrograde tracer FastBlue was injected into male rats’ BLA or entorhinal cortex (EC) 1 week prior to 10 days of immobilization stress. After cessation of stress, FastBlue-labeled and unlabeled IL pyaramidal neurons were loaded with fluorescent dye Lucifer Yellow to visualize dendritic arborization and spine density. As has been previously reported, randomly selected (non-FastBlue-labeled) neurons showed stress-induced dendritic retraction in apical dendrites, an effect also seen in EC-projecting neurons. In contrast, BLA-projecting neurons showed no remodeling with stress, suggesting that this pathway may be particularly resilient against the effects of stress. No neurons showed stress-related changes in spine density, contrasting with reports that more dorsal areas of the mPFC show stress-induced decreases in spine density. Such region- and circuit-specificity in response to stress could contribute to the development of stress-related mental illnesses.
amygdala; chronic stress; connectivity; infralimbic cortex; neural plasticity; spine density
Understanding effects of estrogen on the medial prefrontal cortex (PFC) may help to elucidate the increased prevalence of depression and post-traumatic stress disorder in women of ovarian cycling age. Estrogen replacement in ovariectomized (OVX) young rats amplifies the detrimental effects of stress on working memory (a PFC-mediated task), but the mechanisms by which this occurs have yet to be identified. In male rats, stimulation of norepinephrine alpha-2 adrenoceptors protects working memory from stress-induced impairments. However, this effect has not been studied in females, and has not been examined for sensitivity to estrogen. The current study asked whether OVX females with estrogen replacement (OVX + Est) and without replacement (OVX + Veh) responded differently to stimulation of alpha-2 adrenoceptors after administration of the benzodiazepine inverse agonist FG7142, a pharmacological stressor. The alpha-2 agonist, guanfacine, protected working memory from the impairing effects of FG7142 in OVX + Veh, but not in OVX + Est rats. Western Blot analysis for alpha-2 receptors was performed on PFC tissue from each group, but no changes in expression were found, indicating that the behavioral effects observed were likely not due to changes in receptor expression. These findings point to possible mechanisms by which estrogen may enhance the stress response, and hold implications for the gender discrepancy in the prevalence of stress-related mental illness.
Acute stress; estrogen; norepinephrine; prefrontal cortex; sex differences; working memory
The encoding of reward-predictive stimuli by neurons in the nucleus accumbens (NAcc) depends on integrated synaptic activity from the basolateral amygdala (BLA) and medial prefrontal cortex (mPFC) afferent inputs. In a previous study, we found that single electrical stimulation pulses applied to the BLA facilitate mPFC-evoked spiking in NAcc neurons in a timing-dependent manner, presumably by a fast glutamatergic mechanism. In the present study, the ability of repetitive BLA activation to modulate synaptic inputs to NAcc neurons through dopamine- or N-methyl-d-aspartate (NMDA)-dependent mechanisms is characterized. NAcc neurons receiving excitatory input from both mPFC and BLA were recorded in urethane-anesthetized rats. Train stimulation of the BLA depressed mPFC-evoked spiking in these neurons. This was not attributable to mechanisms involving NMDA or dopamine D1, D2, D3 or D5 receptors, since blockade of these receptors did not affect the BLA-mediated depression. BLA-mediated depression was only evident when the BLA stimulation evoked spikes in the recorded neuron; thus, depolarization of the recorded neuron may be critical for this effect. The ability of the BLA to suppress mPFC-to-NAcc signaling may be a mechanism by which normal or pathologically heightened emotional states disrupt goal-directed behavior in favor of emotionally-driven responses.
electrophysiology; in vivo; heterosynaptic; limbic; reward; addiction
Medial prefrontal cortex (mPFC) neurons respond to Pavlovian conditioned stimuli, and these responses depend on input from the basolateral amygdala (BLA). In this study, we examined the mPFC efferent circuits mediating conditioned responding by testing whether specific subsets of mPFC projection neurons receive BLA input and respond to conditioned stimuli. In urethane-anesthetized rats, we identified mPFC neurons that projected to the nucleus accumbens (NAcc) or to the contralateral mPFC (cmPFC) using antidromic activation. Stimulation of the BLA and Pavlovian conditioned odors selectively activated a subpopulation of ventral mPFC neurons that projected to NAcc, but elicited virtually no activation in mPFC neurons that projected to cmPFC. BLA stimulation typically evoked inhibitory responses among nonactivated neurons projecting to either site. These results suggest that the ventral mPFC-to-NAcc pathway may support behavioral responses to conditioned cues. Furthermore, because projections from the BLA (which also encode affective information) and the mPFC converge within the NAcc, the BLA may recruit the mPFC to drive specific sets of NAcc neurons, and thereby exert control over prefrontal cortical-striato-thalamocortical information flow.
corticostriatal; electrophysiology; emotion; in vivo; infralimbic; fear
Medial prefrontal cortex (mPFC) neurons respond to Pavlovian conditioned stimuli, and these responses depend on input from the basolateral amygdala (BLA). In this study, we examined the mPFC efferent circuits mediating conditioned responding by testing whether specific subsets of mPFC projection neurons receive BLA input and respond to conditioned stimuli. In urethane-anesthetized rats, we identified mPFC neurons that projected to the nucleus accumbens (NAcc) or to the contralateral mPFC (cmPFC) using antidromic activation. Stimulation of the BLA and Pavlovian conditioned odors selectively activated a subpopulation of ventral mPFC neurons that projected to NAcc, but elicited virtually no activation in mPFC neurons that projected to cmPFC. BLA stimulation typically evoked inhibitory responses among non-activated neurons projecting to either site. These results suggest that the ventral mPFC-to-NAcc pathway may support behavioral responses to conditioned cues. Furthermore, because projections from the BLA (which also encode affective information) and the mPFC converge within the NAcc, the BLA may recruit the mPFC to drive specific sets of NAcc neurons, and thereby exert control over prefrontal cortical-striato-thalamocortical information flow.
corticostriatal; electrophysiology; emotion; in vivo; infralimbic; fear
The development of sensitization to amphetamine (AMPH) is dependent on increases in excitatory outflow from the medial prefrontal cortex (mPFC) to subcortical centers. These projections are clearly important for the progressive enhancement of the behavioral response during drug administration that persists through withdrawal.
The objective of this study was to identify the mPFC subcortical pathway(s) activated by a sensitizing regimen of AMPH.
Using retrograde labeling techniques, Fos activation was evaluated in the predominant projection pathways of the mPFC of sensitized rats following a challenge injection of AMPH.
There was a significant increase in Fos-immunoreactive cells in the mPFC, nucleus accumbens (NAc), basolateral amygdala (BLA), and lateral hypothalamus (LH) of rats treated repeatedly with AMPH when compared to vehicle-treated controls. The mPFC pyramidal neurons that project to the LH, but not the NAc or BLA, show a significant induction of Fos after repeated AMPH treatment. In addition, we found a dramatic increase in Fos-activated orexin neurons.
The LH, a region implicated in natural and drug reward processes, may play a role in the development and persistence of sensitization to repeated AMPH through its connections with the mPFC and possibly through its orexin neurons.
dopamine; psychostimulant; behavioral sensitization; prelimbic cortex; infralimbic cortex; reward; addiction; Fluoro-Gold; orexin
The neural circuit linking the medial prefrontal cortex (mPFC) and the basolateral amygdala (BLA) has crucial roles in both the acquisition and the extinction of fear. However, the mechanism by which this circuit encodes fear and extinction remains unknown. In this study, we monitored changes in the magnitude of evoked field potentials (EFPs) in the mPFC–BLA and BLA–mPFC pathways following auditory fear conditioning and extinction, in freely moving rats. We report that extinction of fear is mediated by depression of the EFPs in the mPFC–BLA and by potentiation in the reciprocal pathway of BLA–mPFC. Interestingly, reinstatement of fear was associated with recovery of freezing and with reversal of the changes in EFPs that were observed following extinction in both pathways. The findings indicate that the mPFC–BLA circuit expresses differential changes following fear and extinction and point to dynamic and plastic changes underlying fear, extinction, and reinstatement. Manipulations targeting these different types of plasticity could constitute a therapeutic tool for the treatment of anxiety disorders.
fear; extinction; reinstatement; mPFC; BLA; learning and memory; plasticity; biological psychiatry; psychiatry and behavioral sciences; fear; prefrontal; extinction; amygdale
Reward-seeking behavior is controlled by neuronal circuits that include the basolateral nucleus of amygdala (BLA), medial prefrontal cortex (mPFC), nucleus accumbens (NAc) and ventral tegmental area. Using a discriminative stimulus (DS) task in which an intermittently presented cue (DS) directs the animals to make an operant response for sucrose, we previously demonstrated that dopamine receptor antagonism in the NAc reduced reinforced cue responding, whereas general inactivation of the NAc increased behavioral responding in the absence of the cue. Because they send major glutamatergic projections to the NAc, the BLA and mPFC may also contribute to reward-seeking behaviors modulated by the NAc. In this study we compare the effects of BLA and mPFC inactivation on the performance of a DS task. BLA inactivation by combined GABAA and GABAB agonists impaired cue responding with minimal effects on operant behavior in the absence of cues. Dorsal mPFC (dmPFC) inactivation also inhibited cue-evoked reward-seeking. In contrast, ventral mPFC (vmPFC) inactivation disinhibited responding to unrewarded cues with less influence on reinforced cue responding. These findings demonstrate that the BLA and dmPFC facilitate cue-evoked reward-seeking, whereas, in the same task the vmPFC exerts inhibitory control over unrewarded behaviors.
basolateral nucleus; infralimbic cortex; nucleus accumbens; prelimbic cortex; cingulate cortex; reward-seeking behavior
Cognitive deficits such as impaired decision-making can be a consequence of persistent pain. Normal functions of the intact amygdala and prefrontal cortex are required for emotion-based decision-making that relies on the ability to assess risk, attribute value and identify advantageous strategies. We tested the hypothesis that pain-related cognitive deficits result from amygdala-driven impairment of medial prefrontal cortical (mPFC) function. To do this, we used electrophysiological single-unit recordings in vivo, patch-clamp in brain slices, and various behavioral assays to show that increased neuronal activity in the amygdala in an animal model of arthritis pain was accompanied by decreased mPFC activation and impaired decision-making. Further, pharmacologic inhibition (with a CRF1 receptor antagonist) of pain-related hyperactivity in the basolateral amygdala (BLA), but not central amygdala (CeA), reversed deactivation of mPFC pyramidal cells and improved decision-making deficits. Pain-related cortical deactivation resulted from a shift of balance between inhibitory and excitatory synaptic transmission. Direct excitatory transmission to mPFC pyramidal cells did not change in the pain model whereas polysynaptic inhibitory transmission increased. GABAergic transmission was reduced by non-NMDA receptor antagonists, suggesting synaptic inhibition was glutamate-driven. The results are consistent with a model of BLA-driven feed-forward inhibition of mPFC neurons. In contrast to the differential effects of BLA versus CeA hyperactivity on cortical-cognitive functions, both amygdala nuclei modulate emotional-affective pain behavior. Thus this study shows that the amygdala contributes not only to emotional-affective but also cognitive effects of pain. The novel amygdalo-cortical pain mechanism has important implications for our understanding of amygdala functions and amygdalo-cortical interactions.
Amygdala; prefrontal cortex; pain; anxiety; decision-making; central sensitization; synaptic plasticity
Considerable evidence indicates that the basolateral complex of the amygdala (BLA) interacts with efferent brain regions in mediating glucocorticoid effects on memory consolidation. Here, we investigated whether glucocorticoid influences on the consolidation of memory for emotionally arousing training depend on functional interactions between the BLA and the medial prefrontal cortex (mPFC), a brain region involved in higher-order cognitive and affective processing. The glucocorticoid receptor (GR) agonist RU 28362 administered unilaterally into the left mPFC of male Sprague-Dawley rats immediately after inhibitory avoidance training enhanced 48-h retention performance. An ipsilateral, but not contralateral, lesion of the BLA blocked the memory enhancement. In a second experiment RU 28362 infused into the mPFC after inhibitory avoidance training increased BLA levels of phosphorylated extracellular signal-regulated kinase 1/2 (pErk1/2). Blockade of this pErk1/2 activity in the BLA with the mitogen-activated kinase protein-Erk kinase inhibitor PD98059 prevented the memory enhancement, suggesting that GR agonist administration into the mPFC enhances memory consolidation via modulation of BLA activity. Conversely, GR agonist infusions administered into the BLA posttraining increased pErk1/2 levels in the mPFC in regulating memory consolidation. Moreover, as assessed with a two-phase inhibitory avoidance procedure designed to separate modulatory influences on memory of context and footshock, posttraining GR agonist infusions into either the BLA or mPFC enhanced memory of the contextual as well as aversive information acquired during inhibitory avoidance training. These findings indicate that glucocorticoid effects on memory consolidation depend on bidirectional interactions between the BLA and mPFC.
GR agonist; RU 28362; pErk; MAP kinase; inhibitory avoidance; Glucocorticoid
Anatomical alterations in the medial prefrontal cortex (mPFC) are associated with hypothalamo-pituitary adrenal (HPA) axis dysregulation, altered stress hormone levels, and psychiatric symptoms of stress-related mental illnesses. Functional imaging studies reveal impairment and shrinkage of the mPFC in such conditions, and these findings are paralleled by experimental studies showing dendritic retraction and spine loss following repeated stress in rodents. Here we extend this characterization to how repeated stress affects dendritic spine morphology in mPFC through the utilization of an automated approach which rapidly digitizes, reconstructs 3-dimensionally, and calculates geometric features of neurons. Rats were perfused after being subjected to 3 weeks of daily restraint stress (6 hours/day), and intracellular injections of Lucifer Yellow were made in layers II/III pyramidal neurons in the dorsal mPFC. To reveal spines in all angles of orientation, deconvolved high-resolution confocal laser scanning microscopy image stacks of dendritic segments were reconstructed and analyzed for spine volume, surface area, and length using a Rayburst-based automated approach (8,091 and 8,987 spines for control and stress, respectively). We found that repeated stress results in an overall decrease in mean dendritic spine volume and surface area, which was most pronounced in the distal portion of apical dendritic fields. Moreover, we observed an overall shift in the population of spines, manifested by a reduction in large spines and increase in small spines. These results suggest a failure of spines to mature and stabilize following repeated stress, and are likely to have major repercussions on function, receptor expression, and synaptic efficacy.
dendritic spine; morphometry; plasticity; prefrontal cortex; stress
Recent studies suggest that in female monkeys and rats, estrogens elicit dendritic spine synapse formation in the prefrontal cortex, an area that similar to the hippocampus, plays a critical role in cognition. However, whether gonadal hormones induce synaptic remodeling in the male prefrontal cortex remains unknown. Here we report that gonadectomy reduced, while administration of 5α-dihydrotestosterone or estradiol-benzoate to castrated male rats increased the number of medial prefrontal cortical (mPFC) spine synapses, with estradiol-benzoate being less effective than 5α-dihydrotestosterone. To investigate whether the androgen receptor contributes to the mediation of these changes, we compared the response of testicular feminization mutant (Tfm) male rats to that of wild-type animals. The number of mPFC spine synapses in gonadally intact Tfm rats and 5α-dihydrotestosterone-treated castrated Tfm males was considerably reduced compared to intact wild-type animals, while the synaptogenic effect of estradiol-benzoate was surprisingly enhanced in Tfm rats. These data are consistent with the hypothesis that remodeling of spine synapses in the prefrontal cortex may contribute to the cognitive effect of gonadal steroids. Our findings in Tfm animals indicate that androgen receptors may mediate a large part of the synaptogenic action of androgens in the mPFC of adult males. However, because this effect of 5α-dihydrotestosterone is not completely lost in Tfm rats, additional mechanisms may also be involved.
gonadal hormones; androgen receptor; synaptic plasticity; prefrontal cortex; Tfm rat; stereology
Basolateral amygdala (BLA) and medial prefrontal cortex (mPFC) interactions have been implicated in cue-elicited craving and drug seeking. However, the neurochemical mechanisms underlying drug/environment associations are ill-defined. We used in vivo microdialysis and pharmacological inactivation techniques to identify alterations in mPFC glutamate (GLU) and gamma-aminobutyric acid (GABA) transmission in response to cues previously associated with experimenter-administered cocaine (COC) and the BLA contribution to these effects. Rats received alternate day injections of COC and saline (SAL) paired with a distinct environment for 6 days. Behavioral, neurochemical and immunohistochemical studies were conducted, in drug-free animals, 24 h after the last conditioning session. Animals exposed to a COC-paired environment demonstrated an augmented locomotor activity (LMA) relative to those exposed to the SAL-paired environment. mPFC GABA neurotransmission in the COC-paired environment was significantly increased, whereas GLU overflow was unaltered. Dual labeling of cFos and glutamic acid decarboxylase 67 immunoreactivity in mPFC neurons revealed significantly greater colocalization of these proteins following exposure to the COC-associated environment (CAE) relative to pseudo-conditioned rats or rats exposed to the SAL-associated environment indicating that the conditioned neurochemical response to the COC-paired environment is associated with activation of intrinsic mPFC GABA neurons. BLA inactivation prevented the increase in LMA and the augmentation of mPFC GABA transmission produced by cue exposure. Intra-mPFC application of the AMPA/KA receptor antagonist, NBQX, produced similar effects. These findings indicate that exposure to a CAE increases mPFC GABA transmission by enhancing excitatory drive from the BLA and activation of AMPA/KA receptors on mPFC GABA neurons.
basolateral amygdala; cocaine; drug-associated environment; glutamate; GABA; medial prefrontal cortex; addiction & substance abuse; GABA; glutamate; neurochemistry; cocaine; drug-associated environment; medial prefrontal cortex; basolateral amygdala
When faced with an inescapable stressor, animals may engage in ‘coping’ behaviors, such as chewing inedible objects, that attenuate some physiological responses to the stressor. Previous evidence indicates that dopamine neurotransmission in the right prefrontal cortex is modulated by coping processes. Here we tested whether medial prefrontal cortical (mPFC) neuronal activation, as measured by Fos-immunoreactivity (Fos-ir), was altered in rats chewing inedible objects during exposure to novelty stress. We found that chewing caused an increase in Fos-ir that was selective for the right hemisphere of the mPFC along with a decrease in Fos-ir that was selective for the right central nucleus of the amygdala (CeA), a region that may regulate dopamine neurotransmission in mPFC. These observations suggest that coping during stress engages mPFC and CeA neuronal activity asymmetrically.
stress; hemispheric asymmetry; central nucleus of the amygdala; chewing behavior; dopamine
Chronic stress has been shown in animal models to result in altered dendritic morphology of pyramidal neurons of the medial prefrontal cortex (mPFC). It has been hypothesized that the stress-induced dendritic retractions and spine loss lead to disrupted connectivity that results in stress-induced functional impairment of mPFC. While these alterations were initially viewed as a neurodegenerative event, it has recently been established that stress induced dendritic alterations are reversible if animals are given time to recover from chronic stress. However, whether spine growth accompanies dendritic extension remains to be demonstrated. It is also not known if recovery-phase dendritic extension allows for re-establishment of functional capacity. The goal of this study, therefore, was to characterize the structural and functional effects of chronic stress and recovery on the infralimbic (IL) region of the rat mPFC. We compared neuronal morphology of layer V IL pyramidal neurons from animals subjected to 21 days of chronic restraint stress (CRS) to those that experienced CRS followed by a 21 day recovery period. Layer V pyramidal cell functional capacity was assessed by intra-IL long-term potentiation (LTP) both in the absence and presence of SKF38393, a dopamine receptor partial agonist and a known PFC LTP modulator. We found that stress-induced IL apical dendritic retraction and spine loss co-occur with receptor-mediated impairments to catecholaminergic facilitation of synaptic plasticity. We also found that while post-stress recovery did not reverse distal dendritic retraction, it did result in over-extension of proximal dendritic neuroarchitecture and spine growth as well as a full reversal of CRS-induced impairments to catecholaminergic-mediated synaptic plasticity. Our results support the hypothesis that disease-related PFC dysfunction is a consequence of network disruption secondary to altered structural and functional plasticity and that circuitry reestablishment may underlie elements of recovery. Accordingly, we believe that pharmacological treatments targeted at preventing dendritic retraction and spine loss or encouraging circuitry reestablishment and stabilization may be advantageous in the prevention and treatment of mood and anxiety disorders.
infralimbic; dendritic morphology; dendritic spines; dopamine; long-term potentiation
Restraint stress has been shown to elicit numerous effects on hippocampal function and neuronal morphology, as well as to induce dendritic remodeling in the prefrontal cortex (PFC). However, the effects of acute restraint stress on PFC cognitive function have not been investigated, despite substantial evidence that the PFC malfunctions in many stress-related disorders.
The present study examined the effects of restraint stress on PFC function in both male rats and cycling female rats in either the proestrus (high estrogen) or estrus (low estrogen) phase of the estrus cycle. Animals were restrained for 60 or 120 minutes and then tested on spatial delayed alternation, a PFC-mediated task. Performance after stress was compared to performance on a different day under no-stress conditions, and analyzed using analysis of variance (ANOVA).
Sixty minutes of restraint impaired only females in proestrus, while 120 minutes of restraint produced significant impairments in all animals. Increases in task completion times did not affect performance.
These results demonstrate an interaction between hormonal status and cognitive response to stress in female rats, with high estrogen levels being associated with amplified sensitivity to stress. This effect has been previously observed after administration of a pharmacological stressor (the benzodiazepine inverse agonist FG7142), and results from both studies may be relevant to the increased prevalence of stress-related disorders, such as major depressive disorder, in cycling women. Overall, the results show that restraint stress has important effects on the cognitive functions of the PFC, and that hormonal influences in the PFC are an important area for future research.
Pyramidal neurons in the prefrontal cortex (PFC) are important for the control of cognitive and emotional behavior. The medial PFC (mPFC) receives diverse long-range excitatory inputs from the midline thalamus, contralateral mPFC, basolateral amygdala and ventral hippocampus. While axons from these different regions have distinct distributions in the mPFC, their functional connections at the cellular and subcellular levels remain unknown. Here, we use optogenetics to show that layer 2 pyramidal neurons in acute slices of the mouse mPFC receive excitatory inputs from each of these regions. Using a combination of optogenetics and two-photon microscopy, we then determine the subcellular properties of these inputs. We find that different types of inputs make selective contacts at the levels of both dendrites and spines. Using two-photon uncaging, we show that this subcellular targeting strongly influences synaptic efficacy in these neurons. Together, our results show that functional connectivity is finely tuned, with important implications for signal processing in the mPFC.
prefrontal cortex; layer 2 pyramidal neuron; dendrites; spines; synapses; optogenetics; two-photon microscopy; two-photon uncaging
We have previously demonstrated that the basolateral amygdala (BLA) is a key component of a neural circuit mediating memory formation for emotionally relevant stimuli in an ethologically-based model of conditioned fear, termed conditioned defeat (CD). In this model, subjects are socially defeated by a larger, more aggressive hamster. Upon subsequent exposure to a smaller, non-aggressive intruder, the defeated animal will show high levels of submissive behaviors and fail to defend its territory. Here we examined whether the medial prefrontal cortex (mPFC), an area with extensive connections with the amygdala, is also a component of this circuit. Temporary inactivation of the mPFC using muscimol, a GABAA receptor agonist, significantly enhanced the acquisition but not expression of CD, while blockade of GABAA receptors in the mPFC using bicuculline, a GABAA antagonist, impaired acquisition of CD. Given these findings, we next sought to test whether plasticity related to the defeat experience occurs in the mPFC. We infused anisomycin, a protein synthesis inhibitor, in the mPFC but this treatment did not alter the acquisition of CD. In our final experiment, we demonstrated that bicuculline failed to alter the acquisition of CD. Together, these results demonstrate for the first time that while the mPFC is both necessary and sufficient for the acquisition of CD, it does not appear to mediate plasticity related to the defeat experience. In contrast, while plasticity underlying CD does appear to occur in the BLA, GABAergic receptor inhibition in the BLA is not sufficient to enhance CD.
social stress; amygdala; defensive behavior; conditioned fear; aggression
We investigated the effect of estrogen replacement on the structure and function of penetrating brain arterioles (PA) and blood-brain barrier (BBB) permeability.
Female ovariectomized Sprague Dawley rats were replaced with estradiol (E2) and estriol (E3) (OVX+E; N=13) and compared to ovariectomized animals without replacement (OVX; N=14) and intact controls (CTL, proestrous; N=13). Passive and active diameters, percent tone and passive distensibility of pressurized PA were compared. In addition, BBB permeability to Lucifer Yellow, a marker of transcellular transport, was compared in cerebral arteries.
Ovariectomy increased myogenic tone in PA compared to CTL that was not ameliorated by estrogen treatment. Percent tone at 75 mmHg for CTL vs. OVX and OVX+E was 44 ± 3% vs. 51 ± 1% and 54 ± 3% (p<0.01 vs. CTL for both). No differences were found in passive diameters or distensibility between the groups. BBB permeability increased 500% in OVX vs. CTL animals, however, estrogen replacement restored barrier properties: flux of Lucifer Yellow for CTL, OVX and OVX+E was (ng/mL): 3.4 ± 1.2, 20.2 ± 5.3 (p<0.01 vs. CTL) and 6.15 ± 1.2 (n.s.).
These results suggest that estrogen replacement may not be beneficial for small vessel disease in the brain, but may limit BBB disruption and edema under conditions that cause it.
estrogen; brain arterioles; myogenic tone; blood-brain barrier
In the past few decades it has become clear that estrogen signaling plays a much larger role in modulating the cognitive centers of the brain than previously thought possible. We have developed a nonhuman primate (NHP) model to investigate the relationships between estradiol (E) and cognitive aging. Our studies of cyclical E treatment in ovariectomized (OVX) young and aged rhesus monkeys have revealed compelling cognitive and synaptic effects of E in the context of aging. Delayed response (DR), a task that is particularly dependent on integrity of dorsolateral prefrontal cortex (dlPFC) area 46 revealed the following: 1) that young OVX rhesus monkeys perform equally well whether treated with E or vehicle (V), and 2) that aged OVX animals given E perform as well as young adults with or without E, whereas OVX V-treated aged animals display significant DR impairment. We have analyzed the structure of layer III pyramidal cells in area 46 in these same monkeys. We found both age and treatment effects on these neurons that are consistent with behavioral data. Briefly, reconstructions of pyramidal neurons in area 46 from these monkeys showed that cyclical E increased the density of small, thin spines in both young and aged monkeys. However, this effect of E was against a background of age-related loss of small, thin spines, leaving aged V-treated monkeys with a particularly low density of these highly plastic spines and vulnerable to cognitive decline. Our current interpretation is that E not only plays a critically important role in maintaining spine number, but also enables synaptic plasticity through a cyclical increase in small highly plastic spines that may be stabilized in the context of learning. Interestingly, recent studies demonstrate that chronic E is less effective at inducing spinogenesis than cyclical E. We have begun to link certain molecular attributes of excitatory synapses in area 46 to E effects and cognitive performance in these monkeys. Given the importance of synaptic estrogen receptor α (ER-α) in rat hippocampus, we focused our initial studies on synaptic ER-α in area 46. Three key findings have emerged from these studies: 1) synaptic ER-α is present in axospinous synapses in area 46; 2) it is stable across treatment and age groups (which is not the case in rat hippocampus); and 3) the abundance and distribution of synaptic ER-α is a key correlate of individual variation in cognitive performance in certain age and treatment groups. These findings have important implications for the design of hormone treatment strategies for both surgically and naturally menopausal women.
Prefrontal cortex; estrogen; aging; primate; cognition; hormone replacement therapy
Neuropathic pain is a chronic disease resulting from dysfunction within the "pain matrix". The basolateral amygdala (BLA) can modulate cortical functions and interactions between this structure and the medial prefrontal cortex (mPFC) are important for integrating emotionally salient information. In this study, we have investigated the involvement of the transient receptor potential vanilloid type 1 (TRPV1) and the catabolic enzyme fatty acid amide hydrolase (FAAH) in the morphofunctional changes occurring in the pre-limbic/infra-limbic (PL/IL) cortex in neuropathic rats.
The effect of N-arachidonoyl-serotonin (AA-5-HT), a hybrid FAAH inhibitor and TPRV1 channel antagonist, was tested on nociceptive behaviour associated with neuropathic pain as well as on some phenotypic changes occurring on PL/IL cortex pyramidal neurons. Those neurons were identified as belonging to the BLA-mPFC pathway by electrical stimulation of the BLA followed by hind-paw pressoceptive stimulus application. Changes in their spontaneous and evoked activity were studied in sham or spared nerve injury (SNI) rats before or after repeated treatment with AA-5-HT. Consistently with the SNI-induced changes in PL/IL cortex neurons which underwent profound phenotypic reorganization, suggesting a profound imbalance between excitatory and inhibitory responses in the mPFC neurons, we found an increase in extracellular glutamate levels, as well as the up-regulation of FAAH and TRPV1 in the PL/IL cortex of SNI rats. Daily treatment with AA-5-HT restored cortical neuronal activity, normalizing the electrophysiological changes associated with the peripheral injury of the sciatic nerve. Finally, a single acute intra-PL/IL cortex microinjection of AA-5-HT transiently decreased allodynia more effectively than URB597 or I-RTX, a selective FAAH inhibitor or a TRPV1 blocker, respectively.
These data suggest a possible involvement of endovanilloids in the cortical plastic changes associated with peripheral nerve injury and indicate that therapies able to normalize endovanilloid transmission may prove useful in ameliorating the symptoms and central sequelae associated with neuropathic pain.
A dysregulation in central serotonin neurotransmission and omega-3 fatty acid deficiency have been implicated in the pathophysiology of major depression. To determine the effects of omega-3 fatty acid deficiency on indices of serotonin neurotransmission in the adult rat brain, female rats were fed diets with or without the omega-3 fatty acid precursor α-linolenic acid (ALA) during perinatal (E0–P90), post-weaning (P21–P90), and post-pubescent (P60–130) development. Ovariectomized (OVX) rats and OVX rats with cyclic estrogen treatment were also examined. Serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) content, and fatty acid composition were determined in the prefrontal cortex (PFC), and tryptophan hydroxylase-2 (TPH-2), serotonin transporter, and 5-HT1A autoreceptor mRNA expression were determined in the midbrain. ALA deficiency during perinatal (−62%, p=0.0001), post-weaning (−34%, p=0.0001), and post-pubertal (−10%, p=0.0001) development resulted in a graded reduction in adult PFC docosahexaenoic acid (DHA, 22:6n-3) composition. Relative to controls, perinatal DHA-deficient rats exhibited significantly lower PFC 5-HT content (−65%, p=0.001), significant greater 5-HIAA content (+15%, p=0.046), and a significant greater 5-HIAA/5-HT ratio (+73%, p=0.001). Conversely, post-weaning DHA-deficient rats exhibited significantly greater PFC 5-HT content (+12%, p=0.03), no change in 5-HIAA content, and a significantly smaller 5-HIAA/5-HT ratio (−9%, p=0.01). Post-pubertal DHA-deficient and OXV rats did not exhibit significant alterations in PFC 5-HT or 5-HIAA content. Only perinatal DHA-deficient rats exhibited a significant reduction in midbrain TPH-2 mRNA expression (−29%, p=0.03). These preclinical data support a causal link between perinatal omega-3 fatty acid deficiency and reduced central serotonin synthesis in adult female rats that is independent of ovarian hormones.
Omega-3 fatty acids; docosahexaenoic acid (DHA); 5-HT; 5-HIAA; estrogen; prefrontal cortex; tryptophan hydroxylase-2; serotonin transporter; 5-HT1A; female; rat
The medial prefrontal cortex (mPFC) has been implicated as a site of dysfunction and abnormal morphology in major depressive disorder and post-traumatic stress disorder, two illnesses that can be brought on by exposure to stress. In animal models, stress has long been shown to induce impairments in tasks known to be mediated by the mPFC, and recent work has demonstrated that chronic stress can lead to morphological changes in mPFC pyramidal cells. This review explores the current literature on stress-induced dendritic remodeling in the mPFC, with particular focus on new findings that illuminate modulators of these effects.
The pathophysiology of osteonecrosis of the jaw (ONJ) is thought to be linked to suppression of intracortical remodeling. Aim of this study was to determine whether mice, which normally do not undergo appreciable amounts of intracortical remodeling, could be stimulated by ovariectomy to remodel within the cortex of the mandible and if bisphosphonates (BPs) would suppress this intracortical remodeling.
Material and Methods
Skeletally mature female C3H mice were either ovariectomized (OVX) or SHAM operated and treated with two intravenous doses of zoledronic acid (ZOL, 0.06 mg/kg body weight) or vehicle (VEH). This ZOL dose corresponds to the dose given to cancer patients on a mg/kg basis, adjusted for body weight. Calcein was administered prior to sacrifice to label active formation sites. Dynamic histomorphometry of the mandible and femur were performed.
Vehicle-treated OVX animals had significantly higher (8-fold) intracortical remodeling of the alveolar portion of the mandible compared to sham – this was significantly suppressed by ZOL treatment. At all skeletal sites, overall bone formation rate (BFR) was lower with ZOL treatment compared to the corresponding VEH group.
Under normal conditions the level of intracortical remodeling in the mouse mandible is minimal but in C3H mice can be stimulated to appreciable levels with ovariectomy. Based on this, if the suppression of intracortical remodeling is found to be part of the pathophysiology of ONJ, the ovariectomized C3H mouse could serve as a useful tool for studying this condition.
BRONJ; jaw necrosis; osteonecrosis; ONJ; zoledronic acid
Relapse to maladaptive eating habits during dieting is often provoked by stress and there is evidence for a role of ovarian hormones in stress responses and feeding. We studied the role of these hormones in stress-induced reinstatement of food seeking and medial prefrontal cortex (mPFC) neuronal activation in c-fos-GFP transgenic female rats, which express green fluorescent protein (GFP) in strongly activated neurons. Food-restricted ovariectomized or sham-operated c-fos-GFP rats were trained to lever-press for palatable food pellets. Subsequently, lever-pressing was extinguished and reinstatement of food seeking and mPFC neuronal activation was assessed after injections of the pharmacological stressor yohimbine (0.5–2 mg/kg) or pellet priming (1–4 non-contingent pellets). Estrous cycle effects on reinstatement were also assessed in wild-type rats. Yohimbine- and pellet-priming-induced reinstatement was associated with Fos and GFP induction in mPFC; both reinstatement and neuronal activation were minimally affected by ovarian hormones in both c-fos-GFP and wild-type rats. c-fos-GFP transgenic rats were then used to assess glutamatergic synaptic alterations within activated GFP-positive and non-activated GFP-negative mPFC neurons following yohimbine-induced reinstatement of food seeking. This reinstatement was associated with reduced AMPAR/NMDAR current ratios and increased paired-pulse facilitation in activated GFP-positive but not GFP-negative neurons. Together, while ovarian hormones do not appear to play a role in stress-induced relapse of food seeking in our rat model, this reinstatement was associated with unique synaptic alterations in strongly activated mPFC neurons. Our paper introduces the c-fos-GFP transgenic rat as a new tool to study unique synaptic changes in activated neurons during behavior.
Diet; c-fos-GFP transgenic rats; Medial prefrontal cortex; Ovarian hormones; Reinstatement; Relapse; Stress; Yohimbine