RNA from rat dorsal root ganglia was analyzed in search of potentially beneficial cytokines that are induced in dorsal root ganglia by nerve injury. By reverse transcription, the PCR, and Southern blotting, interleukin-6 mRNA was detected during development but not in normal adult dorsal root ganglia, reappeared within 1 d of sciatic nerve transection, was maximally increased after 2 and 4 d, and decreased below the threshold of detection within 1 week. By RNase protection assay, interleukin-6 mRNA was consistently present in RNA from dorsal root ganglia removed from rats 4 d following transection but not in control dorsal root ganglia. Interleukin-6 bioactivity was also present in dorsal root ganglia following nerve injury. By in situ hybridization, interleukin-6 mRNA was localized within large and medium- sized axotomized neurons. In summary, some sensory neurons respond to axotomy with a brisk transient increase in synthesis of interleukin-6. Injury to the sciatic nerve also induced mRNAs for interleukin-1 beta and tumor necrosis factor-alpha in dorsal root ganglia. The inductions of interleukin-1 beta and tumor necrosis factor-alpha mRNAs were later and more sustained than that of interleukin-6 mRNA. The cellular sources of these two cytokines have not been defined.

In low-density, serum-free cultures of neurons from embryonic rat dorsal root ganglia, interleukin-6 supports the survival of less than one third of the neurons yet virtually all of them bear interleukin-6 alpha-receptors. A finding that might explain this selectivity is that interleukin-6 acts on sensory neurons in culture through a mechanism requiring endogenous brain-derived neurotrophic factor. Antibodies or a trkB fusion protein that block the biological activity of brain-derived neurotrophic factor synthesized by dorsal root ganglion neurons also block the survival-promoting actions of interleukin-6 on these neurons. Two results indicate that interleukin-6 influences synthesis of brain-derived neurotrophic factor in adult dorsal root ganglion neurons. Intrathecal infusion of interleukin-6 in rats increases the concentration of brain-derived neurotrophic factor mRNA in rat lumbar dorsal root ganglia. The induction of brain-derived neurotrophic factor in dorsal root ganglion neurons that is seen after nerve injury in rats or wild-type mice is severely attenuated in mice with null mutation of the interleukin-6 gene. In brief, the ability of interleukin-6 to support the survival of embryonic sensory neurons in vitro depends upon the presence of endogenous brain-derived neurotrophic factor and the induction of brain-derived neurotrophic factor in injured adult sensory neurons depends upon the presence of endogenous interleukin-6.

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CNTF (ciliary neurotrophic factor), purified from rabbit sciatic nerves by a relatively simple procedure, is bioactive in tissue culture at low picomolar concentration and appears as a doublet on polyacrylamide gel electrophoresis (PAGE). In these nerves, CNTF accounts for more than one-half of the survival-promoting activity on ciliary neurons. The concentration of CNTF in rabbit sciatic nerves is estimated to be 5 nmol/kg, more than 1000 times higher than would seem to be required to support neurons if the neurotrophic factor were homogeneously distributed. With recombinant DNA technology, rat CNTF has been synthesized in Escherichia coli, purified without denaturating agents, and found to be bioactive at a slightly lower concentration than CNTF extracted from rabbit sciatic nerves. After radioiodination, CNTF retains biological activity but is not specifically internalized and retrogradely transported in motor and sensory axons. In peripheral nerves, ciliary neurotrophic factor differs biologically from nerve growth factor (NGF) by its much higher tissue concentration and apparent lack of internalization by peripheral nerve axons.

The formation of somatic neuromuscular junctions in skeletal muscle is regulated by an extracellular matrix protein called agrin. Here, we have examined the expression and localization of agrin during development of the rodent urinary bladder, as a first step to examining its possible role at autonomic neuroeffector junctions in smooth muscle. We have found that agrin is expressed on the surface of developing smooth muscle cells and in the basement membrane underlying the urothelium. More importantly, agrin is progressively concentrated at parasympathetic varicosities during postnatal development and is present at virtually all junctions in mature muscle. Reverse transcription/polymerase chain reaction analysis has shown that pelvic ganglion neurons that innervate the bladder express LN/z8 agrin, whereas bladder smooth muscle expresses LN/z- agrin. Together, these results demonstrate that nerve and/or muscle agrin becomes localized at cholinergic parasympathetic varicosities in smooth muscle, where it could play a role in the maturation of the neuroeffector junction.