Stress Effects on Neuronal Structure: Hippocampus, Amygdala, and Prefrontal Cortex

Abstract

The hippocampus provided the gateway into much of what we have learned about stress and brain structural and functional plasticity, and this initial focus has expanded to other interconnected brain regions, such as the amygdala and prefrontal cortex. Starting with the discovery of adrenal steroid, and later, estrogen receptors in the hippocampal formation, and subsequent discovery of dendritic and spine synapse remodeling and neurogenesis in the dentate gyrus, mechanistic studies have revealed both genomic and rapid non-genomic actions of circulating steroid hormones in the brain. Many of these actions occur epigenetically and result in ever-changing patterns of gene expression, in which there are important sex differences that need further exploration. Moreover, glucocorticoid and estrogen actions occur synergistically with an increasing number of cellular mediators that help determine the qualitative nature of the response. The hippocampus has also been a gateway to understanding lasting epigenetic effects of early-life experiences. These findings in animal models have resulted in translation to the human brain and have helped change thinking about the nature of brain malfunction in psychiatric disorders and during aging, as well as the mechanisms of the effects of early-life adversity on the brain and the body.

Figure 1

The trisynaptic organization of the hippocampus showing input from the entorhinal cortex to both CA3 and dentate gyrus (DG), with feed forward and feedback connections between these two regions that promotes memory formation in space and time, but at the same time, makes the CA3 vulnerable to seizure-induced excitation (McEwen, 1999). Chronic stress causes apical dendrites of CA3 neurons to debranch and shorten in a reversible manner, and glutamate release by giant mossy fiber terminals is a driving force. Chronic stress also inhibits neurogenesis in DG and can eventually reduce DG neuron number and DG volume (see text).

Figure 2

Giant mossy fiber terminals (MFTs) of DG neurons that synapse in stratum lucidum of CA3 have multiple active sites of glutamate release on the thorny excrescences that penetrate the MFTs. Normally fully packed with synaptic vesicles, the MFTs show depletion of vesicles after 3 weeks of chronic restraint stress (CRS), with the remaining vesicles being near active synaptic zones. As shown at the right, vesicle area is reduced by CRS but packing density of the remaining vesicles is increased and the area occupied by mitochondria is also increased. This suggests that the chronically stressed MFT are not exhausted but rather very active after chronic stress (Magarinos et al, 1997). Interestingly, by microdialysis, glutamate release in hippocampus caused by restraint stress is abolished by adrenalectomy, indicating involvement of adrenal secretions (Lowy et al, 1993). *P<0.001, two-tailed unpaired Student's t-test.

Figure 3

Effect of deep hibernation and induced awakening on the apical dendritic branching density of CA3 pyramidal neurons. (a) Camera lucida drawings of representative CA3 pyramidal neurons from active and hibernating European hamsters. An overlay of concentric rings centered at the cell body was used for Sholl analysis. (b) One-way ANOVA followed by Tukey post hoc test revealed statistical differences among experimental groups between 100 and 180 μm from the soma (hibernation (HIB) vs active and awaken groups, P<0.001) and also between 200 and 260 μm from the soma (HIB vs active and awaken groups (P<0.005). Taken from reference Magarinos et al (2006).

Figure 4

Polysialylate removal from NCAM neural cell adhesion molecule antagonized the effect of chronic immobilization stress (CIS) to shorten dendritic branches of CA3 neurons. Top: (left) Representative camera lucida traces from vehicle and endo-N-treated rats that were exposed to 10 days of CIS. The dendritic arbors were larger in endo-N-treated animals. (right) Excitotoxic challenge causes greater neuron loss in endo-N treated hippocampus compared to saline control. Bottom: (a) CIS shortened the total length of CA3 dendritic arbor, whereas PSA removal produced an increase in these measures. With combination of CIS and endo-N, the total arbor length was shorter than with endo-N alone but still larger than control values. (b) Interestingly, the effects on numbers of branching points followed a similar pattern. (c) Sholl analysis revealed that PSA removal alone produced an elongation of branches located at all distances from the soma (from 0 to 550 μm approximately), whereas CIS alone specifically shortened branches located near the soma (approximately between 100 and 250 μm). Interestingly, PSA removal antagonized this CIS-induced dendrite atrophy. Data are presented as means (±SEM) N=9–10/group. Taken from reference McCall et al (2013).

Figure 5

Basal release of glucocorticoids varies in a diurnal pattern, and release increases several fold after exposure to a stressor. Glucocorticoids can bind with different affinities to glucocorticoid and mineralocorticoid receptors, which are expressed throughout the brain and seem to exist in both membrane-bound form and nuclear form. Adrenal steroids can have both rapid and delayed effects. The effects can result from non-genomic mechanisms (mediated by membrane-associated receptors, see the figure; Karst et al, 2005; Kelly and Levin, 2001), indirect genomic mechanisms (mediated by membrane receptors and second messengers) and genomic mechanisms (mediated by cytoplasmic receptors that move to the nucleus and act as transcription factors; Yamamoto, 1985). Although classical mineralocorticoid and glucocorticoid receptors seem to mediate many of these effects, other membrane-associated receptors, including G-protein-coupled receptors, may also be involved in some of these actions (Orchinik et al, 1992; Tasker et al, 2006). In addition, activated glucocorticoid receptors can translocate to mitochondria and enhance their calcium buffering capacity (Du et al, 2009). Glucocorticoids rapidly induce glutamate release in the hippocampus through a mechanism that is absent when the mineralocorticoid receptor is deleted and that may involve a membrane-associated form of the mineralocorticoid receptor (Karst et al, 2005; Lowy et al, 1993). An indirect way by which glucocorticoids can influence neurotransmission (glutamatergic, as well as GABAergic, cholinergic, noradrenergic, and serotonergic) is through crosstalk with the endocannabinoid system (Katona and Freund, 2008). They rapidly stimulate endocannabinoid production in the brain, whereupon endocannabinoids bind to cannabinoid receptor 1 (CB1) and transient receptor potential cation channel subfamily V member 1 (TRPV1), and inhibit neurotransmitter release (Chavez et al, 2010; Hill and McEwen, 2010). Although a G-protein-coupled receptor is implicated in endocannabinoid production (Di et al, 2009), there is also evidence for a mechanism blocked by Ru486—a selective antagonist of the classical cytoplasmic glucocorticoid receptor—in the rapid actions of glucocorticoids in prefrontal cortex (Hill and McEwen, 2010). Reprinted from reference Popoli et al, 2012 with permission.

Figure 6

Lower mGlu2 in hippocampus is a biomarker of anxious and depressive-like behaviors and response to rapidly acting antidepressants. (a) mGlu2 receptor expression is reduced in the hippocampus and prefrontal cortex of depressed Flinders Sensitive Line (FSL) rats compared with their controls (Flinders Resistant Line, FRL). These changes are rapidly corrected by acetyl-L-carnitine (LAC), whose effects endure for 2 weeks after drug withdrawal. *P<0.05, two-tailed unpaired Student's t-test. (b) Chronic unpredictable stress (CUS) in susceptible individuals results in depressive-like behaviors that are rapidly corrected by LAC (Nasca et al, 2013). Interestingly, only susceptible individuals show reduced mGlu2 protein levels within the hippocampus. ***P<0.001, one-way analysis of variances followed by Tukey's test for the post hoc analysis. (c) The recently introduced screening method using a light–dark chamber allows identification of high (HS) and low (LS) susceptible individuals, which are characterized by baseline differences in anxiety and in the levels of mineralocorticoid receptors (MRs), but not mGlu2 in the hippocampus. When stressed, HS mice show decreased mGlu2 levels in hippocampus and exacerbation of the baseline anxiety-like behavior compared with LS mice, which cope better with stress. These changes are prevented by a single injection of spironolactone (a MR antagonist), but not RU486 (a GR antagonist). **P<0.01, one-way analysis of variances followed by Tukey's test for the post hoc analysis. (d) Acetylation of histone H3K27, which is regulated by P300, is a key mediator of mGlu2 regulation in response to stress and antidepressant treatment. (e) The MR-driven downregulation of mGlu2 expression is summarized in the epigenetic allostasis model, which suggests that individual differences in stress responsivity may originate from unknown epigenetic influences early in life (Nasca et al, 2014).

Figure 7

Potential cellular mechanisms for rapid antidepressant action. (a) Acetyl-L-carnitine (LAC) may act inside and outside the nucleus to promote rapid antidepressant responses. Inside the nucleus, LAC increases mGlu2 transcription by enhancing acetylation of the NF-κB and histone H3K27 bound to mGlu2 promoter gene. Outside the nucleus, LAC may control stability of the neuronal cytoskeleton to regulate dendritic remodeling (Nasca et al, 2013). (b) Graphical representation of known intracellular signaling pathways involved in the regulation of the actin cytoskeleton (adapted from WikiPathways) in which genes altered by acute stress, chronic stress, or after recovery from stress are highlighted (yellow) based on microarray data derived from Gray et al. (2014).

Figure 8

Chronic stress causes remodeling of dendrites and synaptic connections in many brain regions, including not only hippocampus but also amygdala and medial prefrontal and orbitofrontal cortex (top panel). Effects of acute and chronic stress operate in space and time in an inverted U-shaped manner (bottom panel). Acute stress, mediated by glucocorticoids, and excitatory amino acids and other mediators (see Table 1), can enhance excitability and promote memory over minutes to hours as long as the stressor is not overly intense; intense stress can have the opposite effect. Chronic stress causes neuronal remodeling as depicted in top panel in a largely reversible manner, promoting adaptation (eg, increase vigilance and anxiety in a dangerous environment). Yet, if there is no reversal of the stress-induced changes in neuronal architecture, an outside intervention with pharmaceutical agents and behavioral therapies may be needed to correct the imbalance. Finally, seizures, ischemia, and head trauma can trigger uncontrolled activation of excitatory amino acids leading to free radicals and inflammatory tone potentiated by glucocorticoids.

Figure 9

Epigenetics coupled to the glutamatergic gene expression profile in the PFC in response to stress. (a) The tripartite glutamatergic synapse. Adapted and modified by Popoli et al (2012) by permission. (b) Glutamatergic gene expression profile in unstressed age-matched mice and in mice subjected to acute restraint stress within the PFC. Glt-1: principal glutamate transporter, which removes glutamate from the neuronal synaptic cleft into neuroglia and neurons; xCT: the cystine–glutamate exchanger that facilitates glutamate exchange from glial cells to the perisynaptic space; mGluRs: metabotropic glutamate receptors, which mainly inhibit the release of glutamate into the synaptic space and generate excitatory responses; NR1, essential subunit of NMDA receptors: ionotropic glutamate receptors, which mainly regulate mechanisms of synaptic plasticity. Bars represent mean+SEM, significant comparisons with corresponding controls, **P<0.01 (t-test). (c) ROD analysis of H3K27ac immunoreactivity showing the lack of ARS effects in the subregions (ORBm and PrL) of the PFC. (d and e) Cell count of P300-positive cells showing the lack of ARS effects in the layers I–II and III of either the ORBm or PrL subregions of the PFC. Image credit: Allen Institute for Brain Science (e).