H. de Weerd

MICROVASCULAR CHANGES IN AGED RAT FOREBRAIN - EFFECTS OF CHRONIC NIMODIPINE TREATMENT

In the present study the effects of long-term treatment with the 1,4-dihydropyridine calcium antagonist nimodipine on ultrastructural alterations of the microvascular morphology were examined in the frontoparietal cortex, entorhinal cortex and CA1 of the hippocampus in the aged rat. Qualitative observations of cerebral microvasculature of aged (30 months) Wistar rats revealed the presence of microvascular fibrosis, membranous inclusions within the basement membrane and basement membrane thickenings. In several cortical regions the percentage of aberrant microvessels was significantly reduced in the nimodipine-treated rats. The observed microvascular anomalies were classified into five distinct categories of which microvascular fibrosis type II, defined as collagen deposits up to 1 micron within the microvascular basement membrane, showed the strongest reduction in the nimodipine-treated cases. The decrement of the percentage of aberrant microvessels and the relative occurrence of several classes of microvascular deviations showed some variation in the various brain regions examined and was most pronounced in frontoparietal cortex layer III. These results may provide a morphological basis for the improved motor and cognitive performance in aged rats after long-term oral nimodipine administration.

Dendro-dendritic connections between motoneurons in the rat spinal cord: an electron microscopic investigation

The ultrastructural characteristics of identified dendrite bundles in the rat spinal cord were analyzed following retrograde tracing from the soleus muscles. Dendrite bundles are arranged in networks that are heavily interconnected by means of gap junctions. The bundles are formed by at least 10 crossing dendrites travelling in different focal planes. In between dendrites, elongated gap junctional complexes are frequently found. Dendrite lamellar bodies, recently described to occur in relation with gap junctions in the central nervous system were not observed in the present study.

Ultrastructural evidence for direct excitatory retroambiguus projections to cutaneous trunci and abdominal external oblique muscle motoneurons in the cat

The nucleus retroambiguus (NRA) is a group of neurons, located laterally in the caudal medulla oblongata. The NRA is thought to modulate abdominal pressure in the framework of respiration, vomiting, vocalization, probably parturition, and, in all likelihood mating behavior. The NRA exerts this control through its projections to motoneurons to the nucleus ambiguus in the lateral medulla (innervating pharynx, larynx), and spinal cord (innervating cutaneous trunci, intercostal, abdominal, pelvic floor, and lower limb muscles). The nature of these NRA-motoneuronal projections is unknown. In this study we have determined the ultrastructure of the NRA-motoneuronal projections, and especially those to the abdominal external oblique and cutaneous trunci muscles. In four cats 0.1% cholera toxin subunit b was injected in the external oblique and cutaneous trunci muscles to retrogradely label their motoneurons in the spinal cord. Wheat germ agglutinin-conjugated horseradish peroxidase was injected into the NRA to anterogradely label its contralaterally descending fibers to the motoneurons of both muscles. In order to prevent anterograde labeling of ipsilaterally descending systems not originating from the NRA, a hemisection was made at the level of C2 prior to the NRA injection. The ultrastructural results indicate that the majority (60-74%) of the anterogradely labeled NRA-terminals made monosynaptic contacts with retrogradely labeled dendrites of the external oblique and the cutaneous trunci muscle motoneurons. The majority (86-95%) of the NRA terminals made asymmetric synaptic contacts and 79-84% contained round vesicles. These results demonstrate the existence of direct, presumably excitatory, projections from NRA to external oblique and cutaneous trunci muscle motoneurons.

Direct projections from the nucleus retroambiguus to cricothyroid motoneurons in the cat

Vocalization can be elicited by stimulation in the periaqueductal gray (PAG). Light-microscopical tracing and physiological studies have revealed that the PAG uses the nucleus retroambiguus (NRA) as a relay to excite the vocalization muscle motoneurons. Direct NRA projections have been demonstrated to pharyngeal and abdominal wall muscle motoneurons, but not to laryngeal motoneurons. In two cats 0.1% cholera toxin subunit b was injected in the cricothyroid muscle of the larynx to retrogradely label its motoneurons, and 2.5% wheat germ agglutinin-horseradish peroxidase was injected into the NRA to anterogradely label its fibers. The electronmicroscopical results indicate that the NRA fibers make monosynaptic contacts with cricothyroid motoneuronal dendrites. Almost all NRA terminal profiles had asymmetrical synapses and contained mostly round or pleiomorphic vesicles, which strongly suggests that the NRA-cricothyroid motoneuronal projection is an excitatory pathway.

Serotonergic Intraventricular Axons in the habenular region

SummaryBoth intracerebroventricular injection of 5,7-dihydroxytryptamine and electrolytical midbrain-raphe lesions in rats induce degeneration of supraependymal axons (SEAs) normally occurring in large numbers upon the ependyma of the medial habenular nucleus and habenular commissure. It is concluded that the intraventricular axon plexus in the epithalamic region is comprised of serotonergic (5-HT) fibers originating in the dorsal and/or median raphe nuclei. Besides the elimination of SEAs, conspicious features were a marked reduction in the number of cilia, degenerative signs in the habenular ependyma, and the emergence of large numbers of supraependymal macrophages, being most probably involved in phagocytosis of the axonal debris. It is suggested that the nucleus habenulae medialis is influenced serotonergically by the midbrain raphe via (1) a direct projection upon its neurons and (2) an indirect projection by way of the intraventricular axon plexus. The origin of intraventricular macrophages is discussed in relation to recent data in the literature.

Ultrastructural evidence for a direct excitatory pathway from the nucleus retroambiguus to lateral longissimus and quadratus lumborum motoneurons in the female golden hamster.

During mating, the female golden hamster displays a stereotyped specific receptive posture, characterized by lordosis of the back, elevation of the tail, and extension of the legs. Muscles involved in this posture are thought to be iliopsoas, cutaneus trunci, lateral longissimus (LL), and quadratus lumborum (QL). Lesion studies in rats suggest that mating behavior is controlled by the mesencephalic periaqueductal gray (PAG). The PAG does not project directly to the motoneurons innervating the muscles involved in mating, but is thought to make use of the nucleus retroambiguus (NRA) as relay. The NRA is located ventrolaterally in the most caudal medulla, and projects directly to iliopsoas and cutaneus trunci motoneuronal cell groups. The question is whether this is also true for LL and QL muscles. Retrograde HRP tracing experiments revealed that LL and QL motoneurons are located medially in the ventral horn of the T12–L6 and T13–L4 segments, respectively. A subsequent ultrastructural study combined wheatgerm agglutinin‐conjugated horseradish peroxidase injections in the NRA with cholera‐toxin B‐subunit injections in LL and QL muscles. The results revealed monosynaptic contacts between anterogradely labeled NRA‐fiber terminals with retrogradely labeled dendrites of both LL and QL motoneurons. Almost all these terminals had asymmetrical synapses and contained spherical vesicles, suggesting an excitatory function of this NRA‐motoneuronal pathway. These results correspond with the hypothesis that in hamster the PAG‐NRA‐motoneuronal projection not only involves motoneurons of iliopsoas and cutaneus trunci but also of LL and QL. J. Comp. Neurol. 480:352–363, 2004.

Novel Algorithms for Improved Sensitivity in Non-Invasive Prenatal Testing

Non-invasive prenatal testing (NIPT) of cell-free DNA in maternal plasma, which is a mixture of maternal DNA and a low percentage of fetal DNA, can detect fetal aneuploidies using massively parallel sequencing. Because of the low percentage of fetal DNA, methods with high sensitivity and precision are required. However, sequencing variation lowers sensitivity and hampers detection of trisomy samples. Therefore, we have developed three algorithms to improve sensitivity and specificity: the chi-squared-based variation reduction (χ2VR), the regression-based Z-score (RBZ) and the Match QC score. The χ2VR reduces variability in sequence read counts per chromosome between samples, the RBZ allows for more precise trisomy prediction, and the Match QC score shows if the control group used is representative for a specific sample. We compared the performance of χ2VR to that of existing variation reduction algorithms (peak and GC correction) and that of RBZ to trisomy prediction algorithms (standard Z-score, normalized chromosome value and median-absolute-deviation-based Z-score). χ2VR and the RBZ both reduce variability more than existing methods, and thereby increase the sensitivity of the NIPT analysis. We found the optimal combination of algorithms was to use both GC correction and χ2VR for pre-processing and to use RBZ as the trisomy prediction method.