Nanae Fukushima

Differential neurogenesis and gliogenesis by local and migrating neural stem cells in the olfactory bulb

The rostral migratory stream (RMS) is a unique forebrain structure that provides a long-distance migratory route for the neural stem cells of the periventricular region towards the olfactory bulb (OB). The purpose of the study presented here is to examine the extent of neurogenesis and gliogenesis by the neural stem cells of different origins (periventricular vs. intrabulbar) in the OB. After the RMS had been subjected to injury, the rats received intraperitoneal injections of 5-bromodeoxyuridine (BrdU) and were further reared for 2 weeks. Neuronal and glial differentiations of the BrdU(+) cells in the olfactory bulbar granule cell (OB-GCL) and the olfactory glomerular (OB-GL) layers were examined immunohistochemically using antibodies against neuronal (NeuN, neuronal nuclei) and glial (GFAP, glial fibrillary acidic protein) markers in the OBs with injured and uninjured (control) RMS. In the completely RMS-lesioned OB, where migration of the periventricular neural stem cells was inhibited, a small number of BrdU(+) NeuN(+) cells were found in both the OB-GCL and OB-GL. The BrdU(+) NeuN(+) cells accounted for a much higher percentage of the BrdU(+) cells on the control side (OB-GCL, 36.7%; OB-GL, 8.8%) than on the completely RMS-lesioned side (OB-GCL, 3.7%; OB-GL, 0.6%). The percentage of the BrdU(+) GFAP(+) cells relative to the BrdU(+) cells did not show any major difference between the control and completely RMS-lesioned sides. This study revealed differences in neurogenesis and gliogenesis between the local and migrating neural stem cells in the OB of the adult rodent.

Continual replacement of newly-generated olfactory neurons in adult rats

It has been known that stem cells do exist in the central nervous system, and adult neurogenesis is continually taking place in the olfactory bulb during life. We report here, with the combined method of autoradiography using (3)H-thymidine and immunohistochemistry for a neuronal marker, that 65.3-76.9% of calretinin-immunoreactive bulbar neurons are replaced during the short period of 6 weeks in the adult rodent. The results indicate that neuronal replacement is a common phenomenon in the olfactory bulb during life.

Functionally essential neuronal population of the facial motor nucleus

Cranial nerve impairment is one of the more serious complications in neurosurgery. Nevertheless, the important question of how many neurons are required for cranial nerve functions remains unanswered. The VIIth cranial nerve (facial nerve) in mice was subjected to graded crush injuries. After assessment of the facial function, the number of uninjured, healthy facial motor neurons was quantified with a retrograde neuronal tracer. We report that normal facial function is preserved if intact neurons account for more than 56% of the control value, while complete facial paralysis occurs if intact neurons are reduced to less than 32% of the control value.

Bony Landmarks of the Anterior Cruciate Ligament Tibial Footprint A Detailed Analysis Comparing 3-Dimensional Computed Tomography Images to Visual and Histological Evaluations

Background: Although the importance of tibial tunnel position for achieving stability after anterior cruciate ligament (ACL) reconstruction was recently recognized, there are fewer detailed reports of the anatomy of the tibial topographic footprint compared with the femoral side. Hypothesis: The ACL tibial footprint has a relationship to bony prominences and surrounding bony landmarks. Study Design: Descriptive laboratory study. Methods: This study consisted of 2 anatomic procedures for the identification of bony prominences that correspond to the ACL tibial footprint and 3 surrounding landmarks: the anterior ridge, lateral groove, and intertubercular fossa. In the first procedure, after computed tomography (CT) was performed on 12 paired, embalmed cadaveric knees, 12 knees were visually observed, while their contralateral knees were histologically observed. Comparisons were made between macroscopic and microscopic findings and 3-dimensional (3D) CT images of these bony landmarks. In the second procedure, the shape of the bony prominence and incidence of their bony landmarks were evaluated from the preoperative CT data of 60 knee joints. Results: In the first procedure, we were able to confirm a bony prominence and all 3 surrounding landmarks by CT in all cases. Visual evaluation confirmed a small bony eminence at the anterior boundary of the ACL. The lateral groove was not confirmed macroscopically. The ACL was not attached to the lateral intercondylar tubercle, ACL tibial ridge, and intertubercular space at the posterior boundary. Histological evaluation confirmed that the anterior ridge and lateral groove were positioned at the anterior and lateral boundaries, respectively. There was no ligament tissue on the intercondylar space corresponding to the intercondylar fossa. In the second investigation, the bony prominence showed 2 morphological patterns: an oval type (58.3%) and a triangular type (41.6%). The 3 bony landmarks, including the anterior ridge, lateral groove, and intertubercular fossa, existed in 96.6%, 100.0%, and 96.6% of the cases, respectively. Conclusion: There is a bony prominence corresponding to the ACL footprint and bony landmarks on the anterior, posterior, and lateral boundaries. Clinical Relevance: The study results may help create an accurate and reproducible tunnel, which is essential for successful ACL reconstruction surgery.

Regenerative process of the facial nerve: rate of regeneration of fibers and their bifurcations.

After the main trunk of the mouse facial nerve was injured by crushing, a fiber tracing method was used to quantify the facial motor neurons that extended regenerating nerve fibers to the specific site of the facial nerve branch. The total number of motor neurons retrogradely labeled with a fluorescent tracer, Fluoro-Gold (FG), were 0 on postsurgical days (PSDs) 1 and 2, 75+/-25 on PSD3, 264+/-21 on PSD4, 378+/-19 on PSD6, 428+/-19 on PSD8, 491+/-13 on PSD12 and 532+/-15 on PSD16. Assuming that the FG-positive neurons (535+/-11) of the control mice represent 100%, the FG-labeled neurons accounted for 0, 14, 49, 71, 80, 92 and 99% on the corresponding days. Two different fluorescent tracers were applied to the different facial nerve branches 16 days after facial nerve injuries. Double-labeled neurons were consistently found in the nerve-crushed facial nucleus (3.2%), and their number increased in the nerve-transected facial nucleus (12.2%). The present study indicates that the regenerating facial nerve consists of heterogeneous nerve fibers with varying growth rates and that excessive axonal branching occurs more frequently in the nerve-transected than in the nerve-crushed injuries.

Cell dynamics of calretinin-immunoreactive neurons in the rostral migratory stream after ibotenate-induced lesions in the forebrain

It is now apparent that adult neurogenesis is taking place during life in the olfactory bulb (OB) of the rodent brain. In the olfactory nervous system, the precursor cells of the subventricular zone are known to continually proliferate, migrate through the rostral migratory stream (RMS) and differentiate into the bulbar neurons. The RMS, consisting of heterogeneous cell populations of the neural and neuronal precursor cells, is the unique forebrain structure that provides a long-distance migratory route for the precursor cells. The present study was undertaken to examine whether neuronal regeneration, focusing on calretinin-immunoreactive (+) cells, may proceed in the RMS following lesions induced by an excitotoxin. Two days after ibotenate injections, massive degeneration of calretinin (+) cells occurred in the RMS and its adjacent forebrains. Thereafter, calretinin (+) cells gradually increased in the RMS and reached above their control value 2 weeks after ibotenate injections. Removal of the OB also produced a marked increase in calretinin (+) cells in the RMS. Autoradiographic experiments using (3)H-thymidine showed that calretinin (+) cells were continually generated in the RMS and underwent neuronal turnover within 8 weeks in a normal condition. The results indicate that, in terms of calretinin (+) cells, neuronal differentiation and replacement is continually taking place within the RMS, and that the RMS is capable of repopulating those cells which were injured by ibotenate.

Calretinin-immunoreactive neurons in rostral migratory stream: neuronal differentiation.

The purpose of the present study was to examine immunohistochemically the expression of nestin and polysialylated neural cell adhesion molecule in calretinin-immunoreactive neurons of the rostral migratory stream, the restricted pathway through which neuronal precursor cells migrate towards the olfactory bulb. Using mirror sectioning, calretinin-immunoreactive neurons of the rostral migratory stream were shown to co-express nestin (20.8%) and polysialylated neural cell adhesion molecule (61.8%). The results show that calretinin-immunoreactive neurons in the rostral migratory stream still express immature neural and neuronal phenotypes.

Neurotrophic effect of hepatocyte growth factor on neonatal facial motor neurons.

Abstract The neurotrophic effect of hepatocyte growth factor (HGF) on axotomized facial motor neurons was examined after local application of HGF to the proximal facial nerve stump of the neonatal rat on post-natal day one (P1). Motor neuron survival was expressed as the neuronal cell count on the injured side as a percentage of that on the noninjured side. Motor neuron survival of the control group was 76% on P3, 54% on P5 and 23% on P8, that of the HGF-treated group 78% on P3, 69% on P5 and 31% on P8, and that of the brain-derived neurotrophic factor (BDNF)-treated group 91% on P5 and 45% on P8. The motor neuron survival rates were then adjusted by deducting the facial motor neurons corresponding to the uninjured retroauricular branch (20%) of the facial nerve. The adjusted values were 70% (P3), 42% (P5) and 4% (P8) for the control group, 72% (P3), 61% (P5) and 14% (P8) for the HGF-treated group, and 88% (P5) and 32% (P8) for the BDNF-treated group. These findings demonstrate that HGF has a neuroprotective effect on injured facial motor neurons and suggest that HGF has neurotrophic properties distinct from those of BDNF.

Effects of hypoglossal and facial nerve injuries on milk-suckling.

Functional roles of the perioral anatomical structures involved in breastfeeding were examined in newborn rat pups in which the hypoglossal (XII) and facial (VII) nerves had been resected at the neonatal stage. The XII nerve controls tongue movement and is comprised of two functionally distinct branches: the medial branch related to protrusion of the tongue and the lateral branch related to its retraction. Newborn rat pups with bilateral resection of either of the XII nerve components (main trunk: XII‐trunk; medial branch: XII‐med; lateral branch: XII‐lat) failed to suckle milk and did not survive. Unilateral XII nerve‐resected neonates showed different milk‐suckling capabilities, which thus resulted in differences in survival rate (XII‐trunk: 38%; XII‐med: 24%; XII‐lat: 92%) and postnatal growth during the postnatal 3 weeks until P21. Unilateral and bilateral resections of the VII nerve innervating the buccolabial musculature produced lowered suckling capabilities and retarded postnatal growth, although all pups showed 100% survival. The results indicate a crucial role of the tongue, especially of protruding muscular elements innervated by the XII‐med nerve, in breastfeeding. The results also indicate differential effects of the VII and XII nerve components on suckling capability, survival, and postnatal growth of newborn rat pups.

Fate of Newly Formed Periglomerular Cells in the Olfactory Bulb

It is well established that olfactory receptor cells are replaced during life. Periglomerular (PG) cells of the olfactory bulb have recently been demonstrated to be produced following proliferation and migration of periventricular neuronal precursor cells even in adulthood. The purpose of the present study was to examine the fate of newly formed PG cells in adult rodents. Using 5-bromodeoxyuridine (BrdU), we carried out a quantitative immunohistochemical analysis of BrdU-positive cells in the bulbar glomerular layer at different survival periods. Each number of BrdU-positive PG cells per 100 olfactory glomeruli was 34.193.3 (1 week), 57.292.7 (2 weeks), 28.094.7 (4 weeks) and 25.991.6 (8 weeks). These results indicate that bulbar PG cells, similar to olfactory receptor cells, are mostly replaced during life, and that the olfactory system is composed of disposable neuronal networks centrally as well as peripherally.It is well established that olfactory receptor cells are replaced during life. Periglomerular (PG) cells of the olfactory bulb have recently been demonstrated to be produced following proliferation and migration of periventricular neuronal precursor cells even in adulthood. The purpose of the present study was to examine the fate of newly formed PG cells in adult rodents. Using 5-bromodeoxyuridine (BrdU), we carried out a quantitative immunohistochemical analysis of BrdU-positive cells in the bulbar glomerular layer at different survival periods. Each number of BrdU-positive PG cells per 100 olfactory glomeruli was 34.1 +/- 3.3 (1 week), 57.2 +/- 2.7 (2 weeks), 28.0 +/- 4.7 (4 weeks) and 25.9 +/- 1.6 (8 weeks). These results indicate that bulbar PG cells, similar to olfactory receptor cells, are mostly replaced during life, and that the olfactory system is composed of disposable neuronal networks centrally as well as peripherally.