Months Of Regeneration Research Articles

Assessing the Magnitude of the Amazonian Forest Blowdowns and Post-Disturbance Recovery Using Landsat-8 and Time Series of PlanetScope Satellite Constellation Data

Blowdown events are a major natural disturbance in the central Amazon Forest, but their impact and subsequent vegetation recovery have been poorly understood. This study aimed to track post-disturbance regeneration after blowdown events in the Amazon Forest. We analyzed 45 blowdown sites identified after September 2020 at Amazonas, Mato Grosso, and Colombia jurisdictions using Landsat-8 and PlanetScope NICFI satellite imagery. Non-photosynthetic vegetation (NPV), green vegetation (GV), and shade fractions were calculated for each image and sensor using spectral mixture analysis in Google Earth Engine. The results showed that PlanetScope NICFI data provided more regular and higher-spatial-resolution observations of blowdown areas than Landsat-8, allowing for more accurate characterization of post-disturbance vegetation recovery. Specifically, NICFI data indicated that just four months after the blowdown event, nearly half of ΔNPV, which represents the difference between the NPV after blowdown and the NPV before blowdown, had disappeared. ΔNPV and GV values recovered to pre-blowdown levels after approximately 15 months of regeneration. Our findings highlight that the precise timing of blowdown detection has huge implications on quantification of the magnitude of damage. Landsat data may miss important changes in signal due to the difficulty of obtaining regular monthly observations. These findings provide valuable insights into vegetation recovery dynamics following blowdown events.

Closing the Gap: Bridging Peripheral Sensory Nerve Defects with a Chitosan-Based Conduit a Randomized Prospective Clinical Trial.

Introduction: If tensionless nerve coaptation is not possible, bridging the resulting peripheral nerve defect with an autologous nerve graft is still the current gold standard. The concept of conduits as an alternative with different materials and architectures, such as autologous vein conduits or bioartificial nerve conduits, could not replace the nerve graft until today. Chitosan, as a relatively new biomaterial, has recently demonstrated exceptional biocompatibility and material stability with neural lineage cells. The purpose of this prospective randomized clinical experiment was to determine the efficacy of chitosan-based nerve conduits in regenerating sensory nerves in the hand. Materials and methods: Forty-seven patients with peripheral nerve defects up to 26 mm distal to the carpal tunnel were randomized to receive either a chitosan conduit or an autologous nerve graft with the latter serving as the control group. Fifteen patients from the conduit group and seven patients from the control group were available for a 12-month follow-up examination. The primary outcome parameter was tactile gnosis measured with two-point discrimination. The secondary outcome parameters were Semmens Weinstein Monofilament Testing, self-assessed pain, and patient satisfaction. Results: Significant improvement (in static two-point discrimination) was observed six months after trauma (10.7 ± 1.2 mm; p < 0.05) for chitosan-based nerve conduits, but no further improvement was observed after 12 months of regeneration (10.9 ± 1.3 mm). After six months and twelve months, the autologous nerve graft demonstrated comparable results to the nerve conduit, with a static two-point discrimination of 11.0 ± 2.0 mm and 7.9 ± 1.1 mm. Semmes Weinstein Filament Testing in the nerve conduit group showed a continuous improvement over the regeneration period by reaching from 3.1 ± 0.3 after three months up to 3.7 ± 0.4 after twelve months. Autologous nerve grafts presented similar results: 3.3 ± 0.4 after three months and 3.7 ± 0.5 after twelve months. Patient satisfaction and self-reported pain levels were similar between the chitosan nerve conduit and nerve graft groups. One patient required revision surgery due to complications associated with the chitosan nerve tube. Conclusion: Chitosan-based nerve conduits are safe and suitable for bridging nerve lesions up to 26 mm in the hand. Tactile gnosis improved significantly during the early regeneration period, and functional outcomes were similar to those obtained with an autologous nerve graft. Thus, chitosan appears to be a sufficient substitute for autologous nerve grafts in the treatment of small nerve defects in the hand.

Impaired bone quality characterized by apatite orientation under stress shielding following fixing of a fracture of the radius with a 3D printed Ti-6Al-4V custom-made bone plate in dogs

Custom-made implants have recently gained attention in veterinary medicine because of their ability to properly fit animal bones having a wide variety of shapes and sizes. The effect of custom-made implants on bone soundness and the regeneration process is not yet clear. We fabricated a 3D printed Ti-6Al-4V custom-made bone plate that fits the shape of the dog radius, and placed it into the radius where an osteotomy had been made. The preferential orientation of the apatite c-axis contributes to the mechanical integrity of the bone and is a reliable measure of bone quality. We determined this parameter as well as the bone shape and bone mineral density (BMD). The bone portion which lies parallel to the bone plate exhibited bone resorption, decreased BMD, and significant degradation of apatite orientation, relative to the portion outside the plate, at 7 months after the operation. This demonstrates the presence of stress shielding in which applied stress is not transmitted to bone due to the insertion of a stiff bone plate. This reduced stress condition clearly influences the bone regeneration process. The apatite orientation in the regenerated site remained different even after 7 months of regeneration, indicating insufficient mechanical function in the regenerated portion. This is the first study in which the apatite orientation and BMD of the radius were evaluated under conditions of stress shielding in dogs. Our results suggest that assessment of bone repair by radiography can indicate the degree of restoration of BMD, but not the apatite orientation.

Microscopical observations on the regenerating tail in the tuatara Sphenodon punctatus indicate a tendency to scarring, but also influence from somatic growth.

The process of tail regeneration in the tuatara (Sphenodon punctatus) is not entirely known. Similarity to and differences from lizard tail regenerations are indicated in the present histological and ultrastructural study. Regeneration is influenced by the animal's age and ambient temperature, but in comparison to that of lizards it is very slow and tends to produce outgrowths that do not reach the length of the original tail. Although microscopically similar to lizard blastemas, the mesenchyme rapidly gives rise to a dense connective tissue that contains few muscle bundles, nerves, and fat cells. The unsegmented cartilaginous tube forming the axial skeleton is not calcified after 5 months of regeneration, but calcification in the inner region of the cartilage, present after 10 months, increases thereafter. Amyelinic and myelinic peripheral nerves are seen within the regenerating tails of 2-3 mm in length and the spinal cord forms an ependymal tube inside a cartilaginous casing. Tissues of the original tail, like muscles, vertebrae and the adipose mass, are largely replaced by dense connective tissue that occupies most of the volume of the new tail at 5 and 10 months of regeneration. It is unknown whether the differentiation of the dense connective tissue is caused by the relatively low temperature that this species lives under or stems from a genetic predisposition toward scarring as with most other amniotes. Increases of muscle and adipose tissues seen in older regenerated tails derive from somatic growth of the new tail in the years following tail loss and not from a rapid regeneration process like that in lizards.

Differential protein abundance during the first month of regeneration of the Caribbean star coral Montastraea cavernosa

It is critical to determine the methods by which coral colonies regenerate tissue lost to physical injury as they provide the physical structure of coral reef systems. To explore regeneration, circular lesions (12 mm diameter × 3 mm depth) were created in the fall of 2014 on 124 Montastraea cavernosa colonies located in the coastal waters of Grenada and Carriacou (10–12 m depth). Coral regeneration was documented at weekly intervals for 28 days. Repeated measures ANOVA on estimated weekly coral regeneration rates showed that island (p = 0.024) and colony colour (p = 0.024) were the only factors significantly affecting lesion regeneration. Mean rate of lesion closure during the first 28 days was approximately 2.8 mm2 d−1. Four identical circular lesions were created on 30 M. cavernosa colonies (Carriacou, 10–12 m depth) in the fall of 2015. One representative lesion created on each coral colony was re-sampled at each of 14, 21, and 32 or 33 days following injury, and coral tissue was flash-frozen. Tissues from 10 normally pigmented brown colonies were selected for proteomic analysis using tandem mass tags. The initial polyp sample, the day 14, and the final samples were used to quantify the difference in protein abundance as the lesions healed. In the tissue samples 6419 peptides were reliably identified, which corresponded to 906 unique proteins. During the first month of regeneration, 111 proteins were differentially abundant (p < 0.05) on at least one timepoint and of these, 11 were associated with regeneration. An additional 14 proteins were also identified that were differentially abundant (p < 0.05) and were associated with inflammation or antioxidant activity. This work demonstrates, for the first time, the differential abundance of proteins associated with regeneration in a scleractinian coral.

Persistent alterations in active and passive electrical membrane properties of regenerated nerve fibers of man and mice.

Excitability of regenerated fibers remains impaired due to changes in both passive cable properties and alterations in the voltage-dependent membrane function. These abnormalities were studied by mathematical modeling in human regenerated nerves and experimental studies in mice. In three adult male patients with surgically repaired complete injuries of peripheral nerves of the arm 22 months-26 years prior to investigation, deviation of excitability measures was explained by a hyperpolarizing shift in the resting membrane potential and an increase in the passive 'Barrett and Barrett' conductance (GBB) bridging the nodal and internodal compartments. These changes were associated with an increase in the 'fast' K(+) conductance and the inward rectifier conductance (GH). Similar changes were found in regenerated mouse tibial motor axons at 1 month after a sciatic crush lesion. During the first 5 months of regeneration, GH showed partial recovery, which paralleled that in GBB. The internodal length remained one-third of normal. Excitability abnormalities could be reversed by the energy-dependent Na(+)/K(+) pump blocker ouabain resulting in membrane depolarization. Stressing the Na(+) pumping system during a strenuous activity protocol triggered partial Wallerian degeneration in regenerated nerves but not in control nerves from age-matched mice. The current data suggest that the nodal voltage-gated ion channel machinery is restored in regenerated axons, although the electrical separation from the internodal compartment remains compromised. Due to the persistent increase in number of nodes, the increased activity-dependent Na(+) influx could lead to hyperactivity of the Na(+)/K(+) pump resulting in membrane hyperpolarization and neurotoxic energy insufficiency during strenuous activity.

Regeneration of Articular Cartilage in Lizard Knee from Resident Stem/Progenitor Cells.

The epiphysis of femur and tibia in the lizard Podarcis muralis can extensively regenerate after injury. The process involves the articular cartilage and metaphyseal (growth) plate after damage. The secondary ossification center present between the articular cartilage and the growth plate is replaced by cartilaginous epiphyses after about one month of regeneration at high temperature. The present study analyzes the origin of the chondrogenic cells from putative stem cells located in the growing centers of the epiphyses. The study is carried out using immunocytochemistry for the detection of 5BrdU-labeled long retaining cells and for the localization of telomerase, an enzyme that indicates stemness. The observations show that putative stem cells retaining 5BrdU and positive for telomerase are present in the superficial articular cartilage and metaphyseal growth plate located in the epiphyses. This observation suggests that these areas represent stem cell niches lasting for most of the lifetime of lizards. In healthy long bones of adult lizards, the addition of new chondrocytes from the stem cells population in the articular cartilage and the metaphyseal growth plate likely allows for slow, continuous longitudinal growth. When the knee is injured in the adult lizard, new populations of chondrocytes actively producing chondroitin sulfate proteoglycan are derived from these stem cells to allow for the formation of completely new cartilaginous epiphyses, possibly anticipating the re-formation of secondary centers in later stages. The study suggests that in this lizard species, the regenerative ability of the epiphyses is a pre-adaptation to the regeneration of the articular cartilage.

Regeneration of Vestibular Horizontal Semicircular Canal Afferents in Pigeons

Spontaneous regeneration of vestibular and auditory receptors and their innervating afferents in birds, reptiles, and amphibians are well known. Here, we produced a complete vestibular receptor loss and epithelial denervation using an ototoxic agent (streptomycin), after which we quantitatively characterized the afferent innervation of the horizontal semicircular canals following completed regeneration. We found that calyx, dimorph, and bouton afferents all regenerate in a manner the recapitulates the epithelial topography of normal birds, but over a slow time course. Similar to previous findings in the vestibular otolith maculae, regeneration occurs according to a three-stage temporal sequence. Bouton afferents regenerate during the first month of regeneration, followed by calyceal-bearing afferents in the second and third months. Calyx afferents were the last to regenerate in the final stage of recovery after 3 mo. We also found that regenerated afferents exhibited terminal morphologies that are significantly smaller, less complex, and innervate fewer receptor cells over smaller epithelial areas than those that develop through normative morphogenesis. These structural fiber changes in afferent innervation correlate to alterations in gaze responses during regeneration, although the exact underlying mechanisms responsible for behavioral changes remain unknown. Plasticity in central vestibular neurons processing motion information seem to be required to explain the observed morphologic and response adaptations observed in regenerating vestibular systems.

Investigation of the regeneration potential of the recurrent laryngeal nerve (RLN) after compression injury, using neuromonitoring

The aim of this study was to investigate the regeneration potential of RLN after the compression of the nerve, without disrupting its continuity, using neuromonitoring. In the first operation, the RLN and nervus vagus of adult Goettingen minipigs were dissected free, and the neuromonitoring parameters (amplitude, threshold and lag time of signal) were measured. Injury of the RLN was induced using a "bulldog" clamp. When the signal was no longer detectable, after the 15 min regeneration phase, the operation was finished. The neuromonitoring studies (see above) were repeated in a second operation 6 months later. (1) After the first operation, acute clamping of the RLN led to a reduction in the amplitude of the neuromonitoring signal; the lag time and the threshold of signal remained. Complete restitution of the signal was observed during the first regeneration phase. Repeated clamping led to complete disappearance of the signal. (2) During the second operation, i.e., after 6 months of regeneration, the neuromonitoring signals of both RLN and nervus vagus were detected in 93% of the GMP. No statistical differences (p = 0.17) were noticed between the amplitude of the RLN before the nerve injury (first operation) and after nerve regeneration (second operation). A significant increase in the lag time (p < 0.0005) was shown for both RLN and nervus vagus. The acute compression of RLN can only be detected by observing the amplitude of the neuromonitoring signal. Restitutio ad integrum is possible after a short clamping period but it is important to preserve the RLN continuity.

Axon mis-targeting in the olfactory bulb during regeneration of olfactory neuroepithelium.

During development, primary olfactory axons typically grow to their topographically correct target zone without extensive remodelling. Similarly, in adults, new axons arising from the normal turnover of sensory neurons essentially project to their target without error. In the present study we have examined axon targeting in the olfactory pathway following extensive chemical ablation of the olfactory neuroepithelium in the P2-tau:LacZ line of mice. These mice express LacZ in the P2 subpopulation of primary olfactory neurons whose axons target topographically fixed glomeruli on the medial and lateral surfaces of the olfactory bulb. Intraperitoneal injections of dichlobenil selectively destroyed the sensory neuroepithelium of the nasal cavity without direct physical insult to the olfactory neuron pathway. Primary olfactory neurons regenerated and LacZ staining revealed the trajectory of the P2 axons. Rather than project solely to their topographically appropriate glomeruli, the regenerating P2 axons now terminated in numerous inappropriate glomeruli which were widely dispersed over the olfactory bulb. While these errors in targeting were refined over time, there was still considerable mis-targeting after four months of regeneration.

Energy content of Ophiocoma echinata (Echinodermata: Ophiuroidea) maintained at different feeding levels during arm regeneration

Individuals of Ophiocoma echinata were collected from Missouri Key (FL, USA) in September, 1993 and divided among four feeding levels (fed daily, fed once a week, fed once every 2 weeks, and not fed). Each individual was induced to autotomize three adjacent arms at the disk edge. Energy content of disk, intact arms, gonads, and stomach was examined after 2 months of regeneration. Arm regeneration was inversely related to food level, while stomach and gonad weight was directly related to food level. The four body components were ordered stomach>disk>gonads>intact arms, and the three organic components were ordered lipid>protein≫carbohydrate with respect to energy differences between feeding treatments.

Regeneration of implanted splenic tissue in the rat: re-innervation is host age-dependent and necessary for tissue development

The loss of spleen may lead to fatal bacterial infections. To prevent this, splenic autotransplantation has been performed in humans and experimental animals. However, there is still controversy about the protective function of this procedure. Since innervation plays an important role in splenic function, we investigated whether splenic regenerates are re-innervated, and whether this depends on the donor and host age. Splenic tissue (30 mg) was implanted into the greater omentum of either young (2 days) or old (12 months) rats, from either young or old syngeneic animals. After 3 months of regeneration, the weight of the regenerates was determined, PGP + nerve fibers were revealed by immunohistology, and subdivided into nerve fibers of sympathetic (TH +, NPY +) or sensory (SP +, CGRP +) origin. In addition, proliferating (Ki-67 proliferation antigen +) and apoptotic cells (TUNEL technique +) were likewise investigated. No innervation of splenic regenerates was observed after implantation into old hosts, correlating with poorly developed splenic compartments. In contrast, almost normal re-innervation occurred in young hosts after implantation of both young and old splenic tissue. These regenerates showed well-developed splenic compartments and a normal number and tissue distribution of proliferating and apoptotic cells. However, after the implantation of young tissue, the final size of splenic regenerates was three times larger (140±30 vs. 40±10 mg). Thus, re-innervation of splenic implants is necessary for their subsequent development. It is determined by host age, whereas the final size of the splenic regenerates is regulated by donor age-dependent factors. This model is useful for studying both the process leading to initial innervation and the consequences of this innervation.

Light and TEM study of nonregenerated and experimentally regenerated scales of Lepisosteus oculatus (Holostei) with particular attention to ganoine formation.

The structure of nonregenerated and experimentally regenerated scales of the holostean fish Lepisosteus oculatus and the events taking place before and during ganoine deposition on the scale surface were studied. The aim of this study was to answer the question of the origin of the ganoine in lepisosteids, the scales of which are devoid of dentine, and to compare them to ganoine formation in polypterid scales and to enamel formation in teeth. Two adult specimens were used and the scale structure was studied using light and transmission electron microscopy. Regeneration was used as an alternative to the lack of developmental stages and to induce ganoine deposition on the scale surface. Nonregenerated scales are composed of a thick, avascular bony plate capped by ganoine that is covered either by the epidermis or by dermal elements. The ganoine surface is separated from the covering soft tissues by an unmineralized layer, the ganoine membrane. During the first 2 months of regeneration, the bony plate forms. It differs from the bony plate of nonregenerated scales only by its large, woven-fibered central region and by the presence of numerous vascular canals. Shortly before ganoine deposition, the osteoblasts cease their activity and an epithelial sheet comes to contact them and spreads on the bony surface. This epithelial sheet is connected to the epidermis by a short epithelial bridge only and is composed of two layers: the inner ganoine epithelium (IGE), in contact with the bone surface and composed of juxtaposed columnar cells that synthesize the ganoine matrix, preganoine; the outer ganoine epithelium (OGE), composed of elongated cells, the surface of which is separated from the overlying dermal space by a basal lamina. Isolated patches of preganoine are deposited by the IGE cells in the upper part of the osteoid matrix of the scale. The interpenetrated preganoine and osteoid matrices constitute an anchorage zone between ganoine and bone. Preganoine patches fuse and a continuous layer of preganoine is progressively synthesized by the IGE cells. Preganoine progressively mineralizes to become ganoine. The processes of ganoine formation are similar to those known for the ganoine in the polypterid scales and to those described for enamel deposition in teeth. Ganoine is enamel.

The regeneration of axons through normal and reversed peripheral nerve grafts

The effect of proximo-distal orientation of peripheral nerve grafts upon axonal regeneration has been investigated using the sciatic nerve of the rat as a model. To test the hypothesis that the presence of nerve branches within a graft will cause misdirection of axons in normally oriented grafts but not in reversed grafts, all grafts studied contained branches. Qualitative electron microscopic examination of graft ultrastructure revealed no differences in nerve structure related to graft orientation. In most normally oriented grafts, branches persisted up to 12 months after surgery. These branches contained axons which terminated at the end of the branch. In all reverse oriented grafts, and in a small number of normally oriented ones, the branches could not be seen after two or more months of regeneration. Axons sprouting outside of the epineurium of the graft caused the branch to be incorporated into the nerve structure. Axon counts in the distal stump of grafted nerves after twelve months recovery revealed that normally oriented grafts with persistent branches led to poorer peripheral regeneration, especially of unmyelinated fibers. The results indicate that regeneration of axons to their peripheral targets may be facilitated by reversing the graft orientation.

Cerebrospinal fluid contacting neurons inside the regenerating caudal spinal cord ofXenopustadpoles

Abstract This electron microscopical analysis of the regenerating tail spinal cord in Xenopus tadpoles reports the presence of cells recognizable as cerebrospinal fluid contacting neurons (CSFCNs) after 13, 24 and 30 days of regeneration. The majority of these cells are localized in the latero‐ventral portion of the central canal, the same location they occupy during normal development. These neuronal elements are derived from ependymal cells, and after one month of regeneration they receive synapses of not‐determined origin. This research stresses that the regeneration of CSFCNs is a common phenomenon in Xenopus, as well as in other low vertebrates, and suggests that tail CSFCNs could be mechanoreceptors.

Axonal pathfinding during the regeneration of the goldfish optic pathway.

Retinal ganglion cells in fish and amphibians regenerate their axons after transection of the optic nerve. Fiber tracing studies during the third month of regeneration show that the axons have reestablished a basically normal fiber order in the two brachia of the optic tract; axons originating in the ventral hemiretina are concentrated in the dorsal brachium, axons from the dorsal hemiretina in the ventral brachium. Attardi and Sperry (Exp. Neurol. 7:46-64, 1963) first suggested that the reestablishment of the fiber order reflects path-finding by the regenerating axons. Recently, however, Becker and Cook (Development 101:323-337, 1987) have claimed that the fiber order observed at later stages of regeneration is due to secondary axonal rearrangements and that the initial brachial choice is random. In order to evaluate whether regenerating axons are capable of navigating in the optic tract and brachia and on the tectum, the present study examined the pathway choices and the morphology of regenerating axons en route to their tectal targets in goldfish. Subsets of axons were labeled at various time intervals (2 to 30 days) following an optic nerve crush, by intraretinal application of the lipophilic fluorescent tracer 1,1-dioctadecyl-3-3-3'-3'-tetramethylcarbocyanine (DiI). After a survival time of 18 to 72 hours (to allow for diffusion of DiI along the axons), the experimental animals were perfused with fixative and their right and left optic pathways (nerve, tract, and tectum) were dissected free and separated at the chiasm. Fluorescently labeled axons were traced in whole-mounted pathways. Pathway choices were examined at the brachial bifurcation where axons from ventral and dorsal hemiretinae normally segregate. DiI was found to label axons reliably up to their growth cones, even at the earliest stages of regrowth. The pathway choices of the axons were nonrandom. The majority of the ventral axons reached the appropriate, dorsal hemitectum through the appropriate dorsal brachium of the tract. Dorsal axons reached the ventral hemitectum mainly through the ventral brachium. This suggests the presence of specific guidance cues, accessible to the regenerating axons. Differences in the complexity of the growth cones of the regenerating axons (simple in the nerve and tectal fiber layer, complex in the tract and the synaptic layer of the tectum) provide further evidence for specific interactions between the regenerating axons and their substrates along the pathway. These results argue that regenerating retinal axons in fish are capable of axonal path-finding.

Properties of standard avian slow muscle grafts following long-term regeneration.

The tonic anterior latissimus dorsi (ALD) of adult pigeons was orthotopically homografted and evaluated after 11 months of regeneration for histological, histochemical, electromyographic (EMG), and mechanical properties. The resting EMG activity of the grafts was lower in amplitude than that of the controls, but showed the tonic pattern typical for these tonic muscles. The control and grafted muscles had a histochemically homogeneous population of fibers with moderate myofibrillar adenosine triphosphatase activity. Succinic dehydrogenase activity was moderate for the control muscles, but low for the grafts. The regenerated muscles had fewer and smaller fibers and had much larger intersynaptic distances. Both the regenerated and the contralateral control muscles were slow contracting and maintained tetanic tension for prolonged periods with direct electrical stimulation. The relaxation was slower in the grafted muscle than in the control. The grafts produced 40% of the maximum tension of the control muscles, but the rate of tension development was similar between the two groups. The results indicate that the tonic properties were regenerated, but the innervation pattern was altered and the grafted muscles did not have normal mature fibers even after long-term regeneration.

Impaired refinement of the regenerated retinotectal projection of the goldfish in stroboscopic light: a quantitative WGA-HRP study.

The retinotectal projection of the goldfish was studied after regeneration of a cut optic nerve in stroboscopic light, constant light or diurnal light, with the lens removed to blur the retinal image. Retrograde transport of wheatgerm agglutinin, conjugated to horseradish peroxidase, from a standard tectal injection site was used to measure the topographic precision of the projection. The dispersion of labelled retinal ganglion cells, which reflects this precision, was assessed by a method based on distance to nearest neighbour. In normal fish treated similarly, these cells are known to be clustered into about 1% of the retinal area. Early in regeneration, however, they are widely dispersed. The projection map then re-acquires its precision over two or three months. In diurnal light, lens ablation had no effect on refinement of the regenerated map. Constant light increased the number of labelled cells but also had no significant effect on the map. But in stroboscopic light with a continuous pseudorandom pattern of flash intervals (average rate 4.8 Hz), much less refinement was seen. Even after 70-98 days of regeneration, labelled cells remained scattered, on average, over 20% of the retinal area. These retinae were indistinguishable by several criteria from those obtained in diurnal light after only 32-39 days. Mislocated axon terminals, which are largely eliminated during the second and third months of regeneration in diurnal light, evidently persist much longer in stroboscopic light that synchronizes ganglion cell activity across the retina. These results, like previous ones obtained by blocking the transmission of activity to the tectum, support a model of map refinement based on correlation in the firing of neighbouring neurons, which may have wide application within the nervous system.

SCANNING ELECTRON MICROSCOPY OF THE REGENERATED SHELL OF THE MARINE ARCHAEOGASTROPOD, TEGULA.

A window was cut in the first body whorl of the marine snail, Tegula, to induce shell regeneration. At various intervals after the shell window was cut, the window with the regenerated material and the shell surrounding it were prepared for scanning electron microscopy. Initial crystal deposition occurred in association with an organic matrix and appeared as small, spindle-shaped crystals formed by the aggregation of needle-like subunits. The spindles were frequently aggregated into stellate clusters that coalesced to form a sheet of mineralized tissue. After about two months of regeneration, dumbbell-shaped crystal aggregates and spherulites were apparent on the surface of the regenerated shell. The regenerated shell assumed a normal structure after at least four months of regeneration. Crystal deposition also occurred on the normal shell bordering the shell window. The crystals assumed several forms, and their orientation appeared to be determined by the microtopography of the underlying shell.

Stretch receptors in regenerated rat muscle

Although muscle spindles are known to be present in regenerated muscles, it has not previously been reported whether they have any electrophysiological activity. In the present study, rat extensor digitorum longus muscles were traumatized so as to cause all muscle fibers to degenerate; the muscle nerves were either left intact to promote subsequent reinnervation or severed to impede reinnervation. After 2–4 months of regeneration, the muscles were subjected to stretch stimuli and sensory activity was recorded electrophysiologically. Many of the muscles contained stretch-sensitive units that behaved like muscle spindles, although the responses were often highly adaptive and somewhat erratic. In general, the responses from muscles with the nerve left intact were more normal than those from muscles in which the nerve had been severed. Silver staining of the same muscles showed that morphologically recognizable muscle spindles were present, but all were abnormal to varying degrees. These results demonstrate that regenerated muscles can regain some degree of sensory activity in addition to motor functions. The prospect of restored sensory activity may be critical in evaluating the efficacy of graft or transplant procedures for human muscles.