Ultrastructural features of the serotonin innervation in adult rat hippocampus: An immunocytochemical description in single and serial thin sections

Abstract

This study was aimed at characterizing the fine-structural features of the normal serotonin (5-HT) innervation in adult rat hippocampus, by means of electron microscopic immunocytochemistry with a polyclonal antiserum against 5-HT-glutaraldehyde-protein conjugate (donated by Michel Geffard, Bordeaux). Two hippocampal sectors were examined, at mid-level along the septo-temporal axis: CA3-a of Ammon's horn and crest of the dentate gyrus (DG-c). A large number of axonal varicosities (terminals) were sampled in single ultrathin sections, to achieve a statistically significant comparison of their size and of their relative frequency of synaptic specialization, junctional targets and juxtaposed elements, between the oriens and the radiatum layer of CA3-a, and the molecular and the polymorph layer of DG-c. In both CA3-a layers, the microenvironment of the immunostained terminals was also compared to that of a population of unlabeled varicosities randomly selected from the same micrographs. Moreover, 57 varicosities from the oriens and the radiatum layer of CA3-a were visualized in a long series of thin sections, allowing for their examination from end to end in 43 instances. As measured in single sections, hippocampal 5-HT varicosities were of comparable diameter (0.57 μm on the average) in the two anatomical sectors and four neuropil layers examined. As extrapolated stereologically to whole varicosities, the proportion making a synaptic membrane specialization (synaptic incidence) ranged from 18 to 33% (average of 24%), without statistically significant differences between the two sectors and four layers. The synaptic incidence determined directly from serial sections of CA3-a (18%) was nearly identical to that extrapolated from single sections (18.1% in the oriens and 19.5% in the radiatum layer). In both CA3-a and DG-c, the 5-HT varicosities showing a junctional complex were slightly larger than their non-junctional counterparts. In CA3-a, only dendritic shafts were targeted by synaptic 5-HT varicosities, whereas in DG-c there were also a few axo-spinous synapses. The microenvironment of CA3-a 5-HT varicosities differed markedly from that of randomly selected unlabeled varicosities, due to its much lower frequency of synaptic targets and higher frequency of juxtaposed axonal varicosities, at least in the radiatum layer. In all four layers examined, other axonal varicosities were indeed the most frequently encountered neuronal element in the immediate vicinity of immunostained 5-HT varicosities. Neurites and dendritic shafts were also common, but dendritic spines (4%) were relatively infrequent. Thus, the normal 5-HT innervation of adult rat hippocampus is predominantly non-junctional (70–80% of its varicosities), and it exhibits comparable intrinsic and relational features in the four neuropil layers of CA3-a and DG-c.