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The adult frog sciatic sensory neurons have been shown to regenerate in vitro. If a crush injury is made at the beginning of culture, regeneration starts after 3.4 days and proceeds at a rate of ∼0.8 mm/day for several days. Two‐dimensional gel electrophoresis was used to study the patterns of radiolabeled, fast axonally transported proteins during the first 7 days of regeneration. Interest was fo
The effects of protein kinase inhibitors on regeneration in vitro of adult frog sciatic sensory axons were tested. Regeneration of crush‐injured nerves for 8 days in serum‐free medium was inhibited by staurosporine (100 nM) and H‐7 (100 μM), which are both known to inhibit protein kinase C. With the use of a compartmented culture system it could be shown that H‐7 exerted both local (outgrowth regi
Labelled, rapidly transported axonal proteins were shown to be released frog adult frog sciatic sensory neurons, regenerating in vitro after a crush injury. The spatial distribution of the transported, released proteins could accurately be resolved by culturing the nerve on nitrocellulose paper, which trapped the released proteins. The release was located to the crush and to the entire outgrowth r
The sensory axons of the adult frog sciatic nerve have earlier been shown to regenerate in vitro. If a local test crush is made at the initiation of culturing, regeneration starts after 3.4 days and proceeds at a rate of about 0.8-0.9 mm/day for several days. In the present experiments regeneration was inhibited by adenosine in a reversible and dose-dependent fashion. Similarly, both an adenosine
The present study showed that insulin (0.01 μg/ml, ≈︂ 2 nM) inhibited [3H]‐thymidine incorporation in support cells, most likely Schwann cells, of the cultured frog sciatic nerve. A 25–35% inhibition took place in regenerating nerve preparations as well as in preparations devoid of neuronal protein synthesis, i.e., in isolated 5 mm nerve segments and in gangliectomized nerves, suggesting that the
Various phenothiazines (thioridazine, trifluoperazine and chlorpromazine) and dibenzazepines (lofepramine, amitriptyline and desipramine) were studied for effects on fast axonal transport (AXT) in vitro in frog sciatic nerves. AXT, measured as the accumulation of (3H) leucine‐labelled proteins in front of a ligature, was inhibited by more then 50% by all the drugs tested at 0.2 mM concentrations.
We used the in vitro regenerating frog sciatic nerve to look for effects of insulin and insulin-like growth factors I and II (IGF-I, IGF-II) on regeneration of sensory axons and on injury induced support cell proliferation in the outgrowth region. In nerves cultured for 11 days, a physiological dose (10 ng/ml, ≈ nM) of insulin or IGF-II increased ganglionic protein synthesis (by 20% and 50%, respe
Explanted preparations of peripheral nerves with attached dorsal root ganglia of adult mammals and amphibia survive for several days in serum-free medium and can be used to study axonal regeneration in vitro. This review outlines the methods which we routinely use and how they may be applied to study different aspects of axonal regeneration. When the peripheral nerves are crushed in vitro, axons r
Podocalyxin (PODXL) is a highly glycosylated and sialylated transmembrane protein that is up-regulated in various types of tumors and whose expression levels positively correlate with tumor grade. We previously found Podxl to be highly expressed in murine tumorigenic neural stem/progenitor cells (NSPs). Here we investigated the effects of elevated Podxl levels in these cells. NSPs overexpressing P
Leukemia inhibitory factor (LIF) is locally up-regulated after peripheral nerve injury and may be involved in the subsequent regeneration. Here, adult mice with or without LIF gene deletions were used to study the role of LIF in regeneration. The results show that axonal regeneration in vitro from dorsal root ganglia (DRGs) was unaffected by LIF deletion. However, segments from wild type mice prom
The effect on axonal outgrowth of inhibition of phospholipase A2 activity was studied in a recently developed in vitro model, where dorsal root ganglia with attached spinal roots and nerve stumps from young adult mice were cultured in an extracellular matrix material (Matrigel). The phospholipase A2 inhibitors 4-bromophenacyl bromide and oleyloxyethyl phosphorylcholine dose-dependently reduced axo
Neuronal death after injury or disease could result from imbalanced cytokine expression. Linomide (LS-2616, quinoline-3-carboxamide), a synthetic immunomodulator with effects on cytokine production, suppresses autoimmune diseases of the nervous system. Here adult mice were pre-treated with 200 mg/kg/day of Linomide for 9 days, after which the sciatic nerves were crushed. After another 10 days of L
Using [γ32P]ATP and 1-dimensional electrophoresis this report shows that a protein kinase is released in the culture medium from adult frog sciatic nerves during regeneration in vitro. The kinase, which phosphorylated serine and to some extent threonine residues, was released from non-neuronal cells. It showed an increased activity during the 3rd to 6th day after injury, coinciding with the injury
Adult mice sensory ganglia were cultured in an extracellular matrix gel. Analyses of extending axons were made 48 h (long-term) or immediately (short-term) after addition of protein kinase inhibitors. Long- and short-term growth was insensitive to protein kinase A/G inhibition by HA-1004. Long-term protein kinase C inhibition by chelerythrine affected only certain, long axons. In the short-term vi
The actions of neurotrophic factors on sensory neurons of the adult nodose ganglion were studied in vitro. The ganglia were explanted in an extracellular matrix-based gel that permitted observation of the growing axons. Neurotrophin-4 (NT-4) was a very efficient stimulator of outgrowth of axons from the nodose ganglion and had almost doubled the outgrowth length when this was analyzed after 2 days
We have used adult mouse superior cervical ganglia (SCG) to study the role of mitogen activated protein (MAP) kinase activity during axonal outgrowth in vitro. An initial peak in activity within the first hours of culture was followed by a substantially higher activity after 1 to 2 days, a time when axons were actively growing. The latter peak is probably a result of both higher levels of protein
Analogous to the death of developing neurones deprived of trophic factors, nerve injury in adult life could lead to nerve cell death by apoptosis. Here the occurrence of apoptotic mouse sciatic sensory neurones after injury was investigated by nick-labelling DNA breaks. A small proportion of the neurones reliably became apoptotic after injury in vivo. The response was strongly amplified when the n
In the present study the role of cAMP for axonal outgrowth and Schwann cell proliferation was studying using the cultured frog sciatic nerve. An intrinsic rise in nerve injury, both in vitro and in vivo. Treatment with 0.1‐1.0μM forskolin, an activator of the cAMP‐generating enzyme adenylyl cyclase, increased the cAMP content up to 13‐fold, but was yet without effect on axonal outgrowth during an
The local synthesis and subsequent retrograde axonal transport of [35S]methionine‐labelled proteins was studied in the in vitro regenerating adult frog sciatic sensory axons. By the use of a three compartment culture system, proteins in the outgrowth region were selectively labelled. After 2 days in culture a rise in TCA‐insoluble radioactivity was detected in the dorsal root ganglia, which could
