Enhancement of Sprouting and Putative Regeneration of Central Noradrenergic Fibers by Morphine

Treatment of newborn rats with 6-hydroxydopa (6-OHDOPA, 60 μg/g IP) increased the levels of norepinephrine in the adult cerebellum and hindbrain. Concurrent treatment with morphine sulfate (20 μg/g IP) potentiated the response to 6-OHDOPA in the cerebellum and pons-medulla. In addition, increased noradrenergic neurite density in 4 week cerebellar cortex (as observed with histofluorescent staining by glyoxylic acid) suggests that neonatal morphine increased the sprouting of noradrenergic neurons in the 6-OHDOPA treated rats

Alterations in Noradrenergic Innervation of the Brain Following Dorsal Bundle Lesions in Neonatal Rats

Several seemingly conflicting sets of data have been reported on the regenerative capacity of central noradrenergic neurons, following transection of the ascending noradrenergic fiber tract in neonatal rats (Iacovitti et al., Dev Brain Res 1: 21-33, 1981; Jonsson and Sachs, Brain Res Bull 9: 641-650, 1982). In order to more fully investigate changes in noradrenergic neurons in the brain after such a transection, rats were lesioned at various times after birth, sometimes in conjunction with administration of the neurotoxin, 6-hydroxydopa (6-OHDOPA). Animals were sacrifced at 7, 10, 14, 28, 42 or 56 days after birth, in order to assess the pattern of noradrenergic neuronal damage, as well as the recovery rate. Dorsal bundle lesions were associated with neocortical and hippocampal hypoinnervation by noradrenergic fibers, and sprouting of a collateral fiber group, with production of noradrenergic hyperinnervation of the cerebellum and pons-medulla. Recovery of the norepinephrine (NE) content to control levels occurred in the neocortex at 8 weeks, when the dorsal bundle was lesioned at birth. When the lesion was produced at a later time (3 days or 5 days after birth), less recovery in the neocortex and hippocampus was found. Histofluorescent fiber number, as observed with a glyoxylic acid method, correlated with NE changes. It appears that 6-OHDOPA (20 μg/g IP) does not modify long-term recovery from a dorsal bundle lesion, when rats are co-treated at 3 days after birth. However, the length of the proximal noradrenergic fiber stump may be an important factor affecting the capacity for recovery from injury. These results suggest that a shorter fiber stump, as produced with a dorsal bundle lesion at the level of the pons, may be associated with a greater degree of recovery from injury. Also, the younger the rat at the time of injury, the greater appears to be the capacity for regeneration. These results demonstrate that regeneration can occur in one part of the brain without modification of a collateral hyperinnervation of a different part of the brain. Therefore, our findings discount a programming of central noradrenergic fibers to express a specific number of nerve terminal arborizations, (i.e., the pruning hypothesis )

Noradrenergic Fiber Sprouting in the Cerebellum

In order to attain a better understanding of the sprouting response of noradrenergic fibers in the central nervous system (CNS), noradrenergic innervation to the cerebellum was observed by the glyoxylic acid method after a variety of manipulations and in a genetic variant of mouse classified as Purkinje cell degeneration (pcd/pcd). It has been found that a midbrain lesion in rats at birth will result in a collateral sprouting response of noradrenergic fibers in the cerebellum at 8 weeks, as indicated by the increased number of histofluorescent fibers observed in the molecular layer of the cerebellar cortex. Another procedure, treatment of neonatal rats with nerve growth factor alone appears to produce a temporary stimulation of noradrenergic fiber growth in the cerebellum, as observed by the histofluorescent method, although the innervation at 6 weeks or later is ultimately unchanged from the control group. In contrast, NGF (500 units) given to rats in combination with 6-hydroxydopa (6-OHDOPA) (60 μg/g IP) at 3 days postbirth produces a hyperinnervation of the cerebellum by noradrenergic fibers by 2 weeks of age and until at least 8 weeks of age. A third procedure, locus coeruleus implantation, was generally unsuccessful using the procedures described, since the implant was usually non-viable after several days. In a few instances where histofluorescent nuclei were found within the implant, there was an abundance of histofluorescent fibers within and adjacent to the implant, with fibers appearing to grow into host cerebellum. In the final procedure, it was noted that the density of noradrenergic input to the molecular layer of the cerebellar cortex was markedly increased in a genetic mutant mouse, classified as Purkinje cell degeneration (pcd/pcd), which is characterized by the absence of Purkinje cells of the cerebellum in adulthood. However, because of the tissue shrinkage that occurs after loss of Purkinje cells during postnatal development, it is unclear as to whether this observation represents hyperinnervation or a normal complement of fibers in a smaller brain space. The above procedures demonstrate the plasticity of noradrenergic fibers in neonatal cerebellum, a brain region that undergoes considerable postnatal development. The cerebellum is thought to be a good site for studying development/ regeneration/sprouting of noradrenergic fibers in particular, and central axonal processes in general

Opiate-Enhanced Toxicity and Noradrenergic Sprouting in Rats Treated With 6-Hydroxydopa

Because endorphin receptor activation alters the function of the central noradrenergic system, opiates may change the regenerative sprouting of neurons in response to adrenergic neurotoxins. To test this hypothesis, newborn rats were treated with several opioids and 6-hydroxydopa (6-OHDOPA) and the development of the noradrenergic system was evaluated. In combination with 6-OHDOPA morphine and naloxone potentiated the development of norepinephrine (NE) levels in the pons-medulla and cerebellum by four weeks of age. β-Endorphin, Leu- and Met-enkephalin and d-Ala2-enkephalinamide produced a similar effect in the pons-medulla. The effect of morphine was partially attenuated by naloxone. Increased cerebellar noradrenergic histofluorescent staining was observed with the morphine + 6-OHDOPA and naloxone + 6-OHDOPA treatments. Both naloxone and morphine decreased NE levels in the pons-medulla of adult rats treated with 6-OHDOPA. These results suggest that opiates and endorphins may enhance sprouting of noradrenergic neurons following neonatal treatment with 6-OHDOPA, by increasing the toxicity of this neurotoxin

Developmental Localization of Noradrenergic Innervation to the Rat Cerebellum Following Neonatal 6-Hydroxydopa and Morphine Treatment

In order to demonstrate the influence of morphine on the developmental localization of regenerated noradrenergic fibers in rat cerebellum, a glyoxylic histofluorescent method and radiometric assay for norepinephrine (NE) were utilized. An initial reduction of NE in the cerebellum after 6-hydroxydopa [6-OHDOPA; 60 µg/g intraperitoneally (i.p.)] was followed by a return to control levels at 3 days, and an elevation above control levels at 7 days. The initial rates of recovery of NE in the cerebellum of the 6-OHDOPA group of rats and the group receiving morphine (20 µg/g i.p.) in combination with 6-OHDOPA were identical up to 7 days. However, by 14 days NE content was further elevated in the cerebellum of the morphine+6-OHDOPA group. Histofluorescent microscopic observations of the cerebellar cortex correlated with the biochemical findings. A reduction in cerebellar NE content at 3 days was associated with a reduction in the number of visible histofluorescent fibers in the cerebellar cortex. By 7 days the relative number of fibers in the 6-OHDOPA groups was similar to that seen in the control group, but by 9 days the relative number of fluorescent fibers in the cerebellar cortex was increased above control. By 13 days there was a further increase in the relative number of fluorescent fibers in the cerebellar cortex of the morphine+6-OHDOPA group, as compared to the group treated with 6-OHDOPA alone. These findings provide an anatomic correlate for recovery of noradrenergic fibers after 6-OHDOPA, and demonstrate an action of morphine in enhancing regenerative sprouting

Striatal Dopamine Turnover and MIF-I

Because of conflicting reports of the actions of the antiparkinsonian agent L-prolyl-L-leucyl-glycine amide (PLG, MIF-I) on the turnover of Striatal dopamine (DA), this process was reinvestigated. In the present series of studies, it was found that neither our MIF-I (200 ng ICV) nor the MIF-I used by Versteeg et al. [25]was effective in altering the rate of decline of endogenous DA in the caudate nucleus of rats pretreated with α-methyl-p-tyrosine (300 mg/kg IP). In addition, our MIF-I (1 mg/kg IP) did not change endogenous dihydroxyphenylacetic acid (DOPAC) or homovanillic acid (HVA) in rat striatum. These studies indicate that MIF-I does not alter the turnover rate of DA in nigrostriatal neurons. It is possible that MIF-I or some substance released by MIF-I acts at a posfsynaptic receptor site