Chronic stress alters synaptic terminal structure in hippocampus

Abstract

Repeated psychosocial or restraint stress causes atrophy of apical dendrites in CA3 pyramidal neurons of the hippocampus, accompanied by specific cognitive deficits in spatial learning and memory. Excitatory amino acids mediate this atrophy together with adrenal steroids and the neurotransmitter serotonin. Because the mossy fibers from dentate granule neurons provide a major excitatory input to the CA3 proximal apical dendrites, we measured ultrastructural parameters associated with the mossy fiber-CA3 synapses in control and 21-day restraint-stressed rats in an effort to find additional morphological consequences of stress that could help elucidate the underlying anatomical as well as cellular and molecular mechanisms. Although mossy fiber terminals of control rats were packed with small, clear synaptic vesicles, terminals from stressed animals showed a marked rearrangement of vesicles, with more densely packed clusters localized in the vicinity of active zones. Moreover, compared with controls, restraint stress increased the area of the mossy fiber terminal occupied by mitochondrial profiles and consequently, a larger, localized energy-generating capacity. A single stress session did not produce these changes either immediately after or the next day following the restraint session. These findings provide a morphological marker of the effects of chronic stress on the hippocampus that points to possible underlying neuroanatomical as well as cellular and molecular mechanisms for the ability of repeated stress to cause structural changes within the hippocampus.

Figure 1

Localization of the hippocampal stratum lucidum area used for electron microscopic studies. (A) Durcupan-embedded blocks containing 100-μm-thick sections of the dorsal hippocampus were trimmed at the level of the CA3 region (see box). (B) Semithin sections (1.5 μm) stained with toluidine blue were used as a guide to further trim the blocks to an area containing the stratum lucidum (SL). (Bar = 17.5 μm.) (C) Camera lucida tracing of a representative pyramidal neuron impregnated with the “single” section Golgi method (13). DG, dentate gyrus, SO, stratum oriens, SP, stratum pyramidale, SL, stratum lucidum, SR stratum radiatum.

Figure 2

Pre- and postsynaptic elements of the mossy fiber–CA3 pyramidal neuron synapse in Golgi-impregnated material (for methodological details see ref. 13). (A) Mossy fiber axons (short arrows) and synaptic boutons en passant (long arrows) within the stratum lucidum. (B) Excrescences or complex spines in the proximal apical dendrites of a CA3 pyramidal neuron (arrows). (Bar = 30 μm.)

Figure 3

Ultrastructure of MFT (mf) in the stratum lucidum of the dorsal hippocampal CA3 region of control (A, B, and C) and restraint-stressed (D, E, and F) rats. (A) Control MFT filled with small clear vesicles are perforated with giant spines of CA3 proximal apical dendrites. (B and C) Higher magnification of two representative terminals from control nonstressed rats densely packed with small clear vesicles. The terminals form asymmetric synaptic contacts (arrows) with dendritic spines. Electron dense contacts with the dendritic shaft (d) are considered to be zones of adherence, (D) After 3 weeks of restraint stress MFT show a depletion of small clear vesicles. (E and F) Higher magnifications of MFT in stressed animals. Notice the compact clusters of vesicles accumulated near active zones. d, Dendritic shaft; mf, MFT; s, dendritic spine. [Bars = 2.5 μm (A and D) and 1 μm (B, C, E, and F).

Figure 4

Effect of 3 weeks of daily restraint stress on vesicle density of MFT expressed as number of vesicles/μm³. The asterisk indicates P < 0.001 compared with controls (two-tailed unpaired Student’s t test). Bars represent means + SEM.