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Thirty male month-old Wistar rats were divided into groups: All groups were submitted to adaptation, familiarization and maximum load carrying test MLCT. After the experimental period, animals were euthanized and the tibial nerve and plantaris muscle removed and prepared for electron transmission and histochemistry. The following significant results were found: These changes made it possible to increase the area occupied by myelinated fibers keeping their quantity and also reduce the interstitial space; and d association of anabolic steroid and ST increased the area of unmyelinated axons and thickness of the myelin sheath.
Compared to ST, both strategies have similar results. However, Schwann cells increased significantly only in this strategy. In the peripheral nerves, there is a reduction in the quantity of myelinated fibers, loss of the interaction between the axon and Schwann cells, division of the myelin sheath, and redundant changes between the myelin and Ranvier nodes Mortelliti et al.
Also, the age-related decay of the neuromuscular system results in decreased strength production and contraction velocity, problems in maintaining physical balance, higher risk of accidents and falls and difficulty in performing daily tasks. Collectively, these results may negatively influence the quality and quantity of life of the elderly Deschenes, There is a non-linear relationship between loss of muscle mass and strength causing a decrease in muscle quality muscle strength per unit of muscle mass; Goodpaster et al.
Changes related to chronological age progression in motoneurons and peripheral nerves appear to play a key role in the decline of motor function Kanda and Hashizume, Exercise and physical training are known to have beneficial effects on the PNS Booth et al. Evidence has shown that physical training potentiates the muscular function of aged humans and animals and induces morphological, biochemical and physiological alterations in spinal motoneurons of young and middle-aged animals Brown et al.
Recently, Shokouhi et al. Nevertheless, data on the effects of strength training on peripheral nerves of the aged are unknown. In general, significant changes in hormone levels are associated with the aging process and sarcopenia Kovacheva et al.
Androgenic anabolic steroids AAS has been studied for the treatment of several conditions, such as severe burns, cachexia of individuals with aids and cancer, severe anemia, osteoporosis, frailty syndrome, among others Basaria et al. During the last decade, many publications have demonstrated a significant dose-dependent increase in strength and muscle mass, myofibers cross-sectional area CSA enlargement, myonuclei and satellite cells aggregation in the muscles of the elderly Bhasin et al.
A recent meta-analysis has shown that the effects of AAS on the lean mass of the elderly are dose-dependent and time-consuming Krause Neto et al. In general, several reviews have discussed the effects of these steroids on the CNS, since the first observations were reported in this tissue Melcangi et al. However, more recent studies have indicated that PNS also synthesizes and metabolizes steroids and can be a target for these molecules. In fact, neuroactive steroids exert important physiological functions in the PNS and act on the glial and neuronal compartments Rodriguez-Waitkus et al.
This study aimed to analyze the effects of strength training associated with the administration of exogenous testosterone in the peripheral nerve and skeletal muscle morphology of aged Wistar rats.
Each group was submitted to a type of procedure, as described below: For all groups, we provide commercial reference food for rats and water ad libitum. Training protocol was performed during the active period of the animal nocturnal. The strength training sessions were done between 11 a. In the next week, maximum loaded carrying capacity tests MLCT were applied using two familiarization sessions. Testing session was conducted until the animal was unable to climb the ladder for at least two consecutive trials failure.
Between each attempt, the animal had an interval of 2 min. From the week following the completion of the MLCT, rodents were conducted to strength training protocol during 15 weeks 45 sessions. In the first session, rats climbed the ladder twice in each of the following workloads: The animal, then, was able to climb the ladder two more times, totaling at the end eight maximum climbs. If the animal could not complete the protocol with eight maximum climbs, the same loads were maintained for the next training session.
Each rodent had a 2-min interval between each climb. Training was performed three times per week on alternate days. Animals were weighted every Tuesday to recalculate the application dosage. TP administration was done by intraperitoneal injection, twice a week Tuesdays and Fridays , starting from the first day of the experiment, in the AC and STA groups.
The duration of treatment was concomitant with the strength training period of the ST and STA groups i. The animals were euthanized using the CO 2 inhalation method, according to the schedule of each experimental group. After euthanasia, tibial nerve and plantaris PL muscle were removed and prepared for electronic transmission and histochemistry analysis.
Tibial nerve was chosen because of its innervation in the predominant muscles of the posterior region of the lower limbs. The PL muscles were chosen according to their predominance of muscle fiber typology and action during the climbing exercise. The PL is glycolytic and predominantly formed of faster contraction myofibers type II.
Thus, an incision was made in the posterior portion of the right knee of the animal up to his ankle to expose the tibial nerve and PL muscle. After the tibial nerve and PL muscles are exposed, cleaned of fat and connective tissue, we quickly prepared them for specific analyses of light and electronic transmission microscopy. After incision in the posterior portion of the right knee, we removed a fragment of approximately 0. Nerve fragment was placed in 2.
Then the material was washed three times with the same buffer solution for 5 min at a time. The fragments remained overnight in 0. In the morning, it was washed with the plug, dehydrated in increasing series of alcohol and propylene oxide for 8 h, under rotation. The nerves were included in the pure resin Spurr , in the position that the nerve fibers were cross-sectioned.
After completion of the material preparation procedure, the histological slides were prepared and the tissue stained with toluidine blue. After selection of the fields in the semifinished sections, the ultra-thin sections were obtained with a diamond knife, in ultramicrotome Sorvall MT-2 , and contrasted with uranyl acetate and lead citrate, finally being analyzed by the electronic transmission microscope.
The material was taken to the electronic transmission microscope Jeol JSM, ICB, USP and photomicrographs of the nerves were captured in cross-shaped fields starting from the top to the bottom of the image and from left to right. In this way, we were able to photograph the material trying to capture the nerve as a whole. Photomicrographs were done using a magnification of and times for stereological and morphometric analysis, respectively.
For the stereological study, we captured 25 images of each group in the transmission electron microscope with a final magnification of times. The following variables were analyzed and quantified:. For the morphometric study, we captured 35 images of each group using a fold increase and performed the following measurements: To measure the CSAs, each structure of interest in the image was surrounded.
Subsequently, the mean diameters were calculated from the mean between largest and smallest diameters of each structure. To calculate the G ratio, we used the quotient between the diameter of the axon and the myelinated fiber. Finally, the mean thickness of the myelin sheath was determined by the average of four equivalent cross-shaped traces on the image. For this, we use the Axiovision 4. Finally, we mounted on Jeli glycerin. Measurements were made in software Axiovision version 4. All data are presented by mean and standard deviation SD.
For comparison of the PRE vs. To compare the same parameters among the trained groups, we used a t -test for independent samples. For the statistical calculations, we used SPSS software version Load per climb g during 15 weeks of experimentation from groups strength trained ST and strength trained plus anabolic steroid STA.
For other analyses, we did not find any significant differences. Nv, numerical density number. Clearly, it is mentioned in the literature that the advance of age affects the function and regeneration of peripheral nerves in both human and experimental models Jeronimo et al. In this study, we did not observe any changes in most of the variables between groups IC and FC.
However, the literature shows a great variability of evidence, citing variations between the types and functions of peripheral nerves, nerve portion, species and rodent lineages analyzed. The authors found no statistical difference in the MF area between the ages of , and days.
However, the myelinated axon area was significantly reduced in the distal segment of the older sample. The explanation may be the effect of aging on the structure of the motoneurons since we know that the number of motoneurons decreases with age Kanda and Hashizume, Nevertheless, there is no clue as to when this process begins or how the process occurs.
Hashizume and Kanda evaluated motor neurons of Fisher rats and found a reduction in the number of motor nuclei of the medial gastrocnemius muscle nerve at the age of 27 months. The authors explained that this may be due to cell death or an increase in the number of unlabeled nuclei. If the axon no longer extends, the axoplasmic transport will not occur, but despite this, such demonstration did not occur to the ulnar nerve.
Furthermore, changes in other components of the PNS, such as a neuromuscular junction, may precede peripheral nerve changes Deschenes et al. This fact brings to light the different effect of aging on the different nerves and their morphology. Recently, Sakita et al. Perhaps, reduction on MF numerical density may precede it. Thus, we can presume that atrophy of these fibers should occur later than expected. The myelin sheath is a fundamental structure in the healthy functioning of healthy nerves Magnaghi et al.
In the present study, we did not find any variation in the thickness of the myelin sheath between IC and FC groups. Our data are corroborated by several studies in the literature. The authors mentioned a gradual increase of myelin sheath lamellae from 1 week of life to 12 months of age. From this point up to 18 months, the myelin sheath remained unchanged. Electrophysiological analysis showed that the amplitude of muscular action potentials gradually increased from 1 week to 18 months of age, showing a positive linear correlation with the number of lamellae in the myelin sheath.
Other studies, whose focus was to evaluate more advanced ages, also showed no changes in the thickness or perimeter of the myelin sheath Hashizume and Kanda, ; Jeronimo et al. However, Ceballos et al. Knowing that the portion of the nerve analyzed can influence the results investigated, Sakita et al.