Manual Tracing Protocol for Rhesus Macaque Hippocampus
The following guidelines detail the anatomical landmarks and procedures used for systematically outlining the hippocampus in the caudal to rostral direction. This protocol is the same as used by Shamy et al. (2006) with minor modifications to accommodate the age range of the present scans (ranging from 1 week to 260 weeks of age).
At its caudal extent, the hippocampus has a roughly a dorsoventral orientation and is primarily encapsulated by white matter. The retrosplenial cortex appears along the medial surface of the hippocampus, separated by a thin band of white matter. White matter also forms the dorsal border of the hippocampus. The hippocampus is bordered laterally by the fornix, and white matter of the temporal lobe forms the ventral surface of the hippocampus.
The outline at this level started at the dorsal-medial extent of the hippocampus and extended laterally to the white matter of the fornix, and extended ventrally from the fornix along the lateral surface of the hippocampus to the white matter ventral to the hippocampus. The outline continued along the white matter to the starting point.
Proceeding rostrally, the dorsal-medial extent of the hippocampus forms a portion of the medial surface of the brain. The pulvinar of the thalamus also appears along the dorsal-medial surface of the hippocampus. The ventral surface of the thalamus is separated from the dorsal surface of the hippocampus by cerebrospinal fluid along the medial surface of the brain. The dorsal-lateral surface of the hippocampus is formed by the fimbria. The lateral border is formed by the temporal horn of the lateral ventricle. The ventral surface of the hippocampus is formed by white matter of the temporal lobe.
The outline at this level began at the medial extent of the hippocampus, which is ventral to the thalamus. It was extended laterally along the dorsal surface of the hippocampus until it reached the fimbria. The outline continued ventrally along the temporal horn of the lateral ventricle to the sub-hippocampus white matter and continued medially along the white matter to the starting point.
In the body of the hippocampus, the medial surface of the hippocampus forms the medial surface of the brain. At this level, white matter of the alveus comprises the dorsal surface of the hippocampus. A small section of the temporal horn of the lateral ventricle may be visible along the dorsal-lateral surface of the hippocampus. The ventral border of the hippocampus is formed by white matter.
The starting point of the outline at this level is the medial extent of the hippocampus on the medial surface of the brain. The outline was drawn laterally along the dorsal surface of the hippocampus, ventral to the alveus, to the temporal horn of the lateral ventricle. The outline continued ventrally and then medially along the white matter of the temporal lobe to the starting point.
Proceeding rostrally, the hippocampus is outlined in much the same way as just described. The starting point for the outline at this level was the medial surface of the hippocampus. The outline was drawn dorsally along the medial surface of the hippocampus to the alveus. The outline continued laterally along the dorsal surface of the hippocampus until it reached either the temporal horn of the lateral ventricle or the white matter of the temporal lobe. The outline was then drawn ventrally along the lateral ventricle or temporal lobe white matter to the sub-hippocampus white matter. It was completed medially along the white matter to the medial surface of the brain.
In rostral sections, the hippocampus forms part of the medial surface of the brain and is bordered dorsally by the alveus, laterally by the temporal horn of the lateral ventricle, and ventrally by white matter of the temporal lobe. When the uncus appears as a separate tissue from the body of the hippocampus, than the two are outlined separately. If there is only fimbria separating the two, the white matter is omitted from the tracing. Otherwise, both the body and uncus are outlines together at this level if connected by grey matter.
The outline was started at the medial surface of the brain and extended laterally along the alveus to the temporal horn of the lateral ventricle. The outline was continued ventrally along the lateral ventricle to the sub-hippocampus white matter. The outline was completed by tracing medially along the white matter to the medial surface of the brain.
At its rostral extent, the hippocampus decreases in size. The entorhinal cortex begins to form part of the medial surface of the hippocampus. The dorsal surface of the hippocampus often appears to be fused with the ventral surface of the amygdala at this level, but the two structures are separated by the alveus or by a thin portion of the lateral ventricle. The hippocampus continues to be bordered laterally by the temporal horn of the lateral ventricle and ventrally by white matter of the temporal lobe.
At this level, the outline began at the dorsal-medial extent of the hippocampus. This point is found at the junction of the alveus and the band of fibers extending from the sub-hippocampus white matter toward the semiannular sulcus. From this point, the outline continued laterally along the alveus, then ventrally along the temporal horn of the lateral ventricle, until the sub-hippocampus white matter was reached. The outline was completed medially along the white matter, which separates the hippocampus from the entorhinal cortex, to the starting point.
Once the hippocampus was traced in the coronal plane, the tracings were verified in the sagittal and axial planes, with specific care taken to specifically exclude amygdala tissue along the amygdalo-hippocampal border at the most rostral aspect of the hippocampus as well as the retrosplenial cortex at the most caudal aspect of the hippocampus.