Progesterone treatment reduces NADPH-diaphorase/nitric oxide synthase in Wobbler mouse motoneuron disease.

Previous work demonstrated that progesterone (PROG) treatment attenuates morphological, molecular and functional abnormalities in the spinal cord of the Wobbler (Wr) mouse, a genetic model of motoneuron degeneration. Wr mice show a marked up-regulation of the nitric oxide synthesizing enzyme (NOS). Since nitric oxide is a highly reactive species, it may play a role in neuropathology of Wr mice. We now studied if PROG neuroprotection involved changes of NOS activity in motoneurons and astrocytes, determined by the nicotinamide adenine dinucleotide phosphate-diaphorase (NADPHD) histochemical reaction. Two and four-month-old Wr mice at the progressive and stabilization stages of the disease, respectively, and their age-matched controls were left untreated or received a single 20-mg PROG pellet for 18 days. PROG reduced the high number of NADPHD-active motoneurons and white matter astrocytes in 2-month-old Wr mice but was unable to change the low number of NADPHD-active motoneurons in 4-month-old Wr mice or astrocytes in this age group. A large number of motoneurons in 2-month-old Wr mice showed a vacuolated phenotype, which was significantly reverted by PROG treatment. In summary, PROG treatment during the early symptomatic stage of the disease caused a significant reduction of NADPHD-active motoneurons and astrocytes and also reduced vacuolated degenerating cells, suggesting that blockade of NO synthesis and oxidative damage may contribute to steroid neuroprotection.

Glucocorticoid effects on Fos immunoreactivity and NADPH-diaphorase histochemical staining following spinal cord injury.

Glucocorticoids (GC) provide neuroprotection and early recovery after spinal cord injury (SCI). While several mechanisms were proposed to account for these effects, limited information exists regarding GC actions in sensory areas of the spinal cord. Presently, we studied the time course of Fos expression, and reduced nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) histochemical staining to monitor neuronal responses to SCI with or without GC treatment. Rats with sham-operation or transection at the thoracic level (T7-T8) received vehicle or 5 mg/kg of the GC dexamethasone (DEX) at 5 min post-lesion and were sacrificed 2 or 4 h after surgery. Another group of SCI rats received vehicle or intensive DEX treatment (5 min, 6 h, 18 h and 46 h post-lesion) and were sacrificed 48 h after surgery. The number of NADPH-d positive neurons or Fos immunoreactive nuclei was studied by computer-assisted image analysis in superficial dorsal horn (Laminae I-III) and central canal area (Lamina X) below the lesion. While constitutive Fos immunoreactive nuclei were sparse in controls, SCI increased Fos expression at 2 and 4 h after injury. DEX treatment significantly enhanced the number of Fos positive nuclei in Laminae I-III by 4 h after transection, although the response was not maintained by intensive steroid treatment when tested at 48 h after SCI. NADPH-d positive neurons in Laminae I-III increased at 2 and 4 h after SCI while a delayed increased was found in central canal area (Lamina X). DEX treatment decreased NADPH-d positive neurons to sham-operated levels at all time points examined. Thus, while GC stimulation of Fos suggests activation of neurons involved in sympathetic outflow and/or pain, down-regulation of NADPH-d indicates attenuation of nociceptive outflow, considering the role of enzyme-derived nitric oxide in pain-related mechanisms. Differential hormonal effects on these molecules agree with their localization in different cell populations.

GMP protects against quinolinic acid-induced loss of NADPH-diaphorase-positive cells in the rat striatum.

When injected into the rat striatum, quinolinic acid causes dose-dependent widespread cell death. All cell types, including the NADPH-diaphorase-positive neurons appear to be sensitive to the toxin. The latter cells are destroyed by quinolinic acid injections of 180 nmol per striatum, this effect being blocked by the concomitant administration of 5 mg/kg of the non-competitive N-methyl-D-aspartate antagonist MK-801. We report that guanosine-5'-monophosphate (GMP), at a dose of 360 nmol, is equally effective in protecting the diaphorase-positive cells against quinolinate toxicity.