Alterations in AMPA receptor subunit expression after experimental spinal cord contusion injury.

The AMPA-preferring subtype of ionotropic glutamate receptors (GluRs) is a hetero-oligomeric ion channel assembled from various combinations of four subunits: GluR1, GluR2, GluR3, and GluR4. Antagonists of these receptors can mitigate the effects of experimental spinal cord injury (SCI), indicating that these receptors play a significant role in pathophysiology after spinal trauma. We tested the hypothesis that SCI alters expression of AMPA receptors using a standardized thoracic weight-drop model of rat contusive spinal cord injury. AMPA receptor subunit expression was measured at 24 hr and at 1 month after SCI with quantitative Western blot analysis and in situ hybridization. GluR2 protein levels were preferentially reduced near the injury site 24 hr after SCI. This reduction persisted at 1 month. At a cellular level, a significant decrease in both GluR2 and GluR4 mRNA was found in spared ventral motor neurons adjacent to the injury site and distal to it, with other AMPA subunit mRNAs maintained at control levels. In contrast, only GluR1 mRNA was decreased in the sympathetic preganglionic neurons of the intermediolateral horn. These results suggest population-specific and long-lasting changes in neuronal AMPA receptor composition, which may alter response to glutamate after SCI. These alterations may contribute not only to acute neuropathological consequences of injury, but they may also be partially responsible for the altered functional state of preserved tissue seen chronically after SCI.

Erythrocyte sodium/potassium adenosine triphosphatase in thyroid disease and nonthyroidal illness.

Thyroid hormone is known to modulate cell membrane sodium/potassium adenosine triphosphatase (Na/K-ATPase). To determine whether the activity of this enzyme differed in patients with nonthyroidal illness with low levels of circulating thyroid hormones and patients with documented clinical hypothyroidism, we measured Na/K-ATPase activity in red blood cells from patients with hypo- and hyperthyroidism, patients with nonthyroid disease with and without reduced circulating levels of thyroid hormone, and normal subjects. We also assessed whether the activity of this enzyme reflects decreased thyroid hormone action at the cellular level in patients with nonthyroidal illness. Hyperthyroidism was associated with decreased and hypothyroidism with increased erythrocyte Na/K-ATPase activity [142 +/- 24 (+/- SE) and 371 +/- 37 nmol Pi/mg X h; P less than 0.05 and P less than 0.01 compared to normal]. Enzyme activity in cells from patients with nonthyroidal illness and low levels of circulating T3 was significantly higher than that in cells from normal subjects (289 +/- 11 vs. 223 +/- 16 nmol Pi/mg X h; P less than 0.01), but was not significantly different from that in cells from hypothyroid patients. Red cell Na/K-ATPase activity in patients with nonthyroidal illness and normal thyroid function tests (185 +/- 38 nmol Pi/mg X h was indistinguishable from normal values. These data confirm that hyperthyroid patients have decreased red cell Na/K-ATPase activity and provide direct evidence that erythrocyte ATPase activity is increased in hypothyroid patients. The change in enzyme activity in patients with nonthyroidal illness and decreased circulating T3 levels was comparable to that in hypothyroidism. These results suggest that since red cell Na/K-ATPase activity does not distinguish between ill patients with low thyroid function tests and those with hypothyroidism, tissue hypothyroidism may exist in the former group of patients.

Pregnancy in women with renal disease and moderate renal insufficiency.

Data were gathered on 25 pregnancies in 23 women with moderate renal insufficiency as defined by a serum creatinine level of 1.4 mg/dl or greater prior to or at the onset of pregnancy. Twelve of the women had primary glomerular diseases, five (accounting for seven pregnancies) had interstitial diseases, and six had other renal diseases. In seven women with baseline serum creatinine levels ranging from 1.7 to 2.7 mg/dl, pregnancy was accompanied by a decline in renal function that was believed to be greater than expected from the natural history of the disease. The change ranged from a rise in serum creatinine level of 1.2 mg/dl to a functional decline that required dialysis. In 14 women (16 pregnancies), renal function either remained stable or declined to a degree consistent with the natural history of the disease. In two, the follow-up period was not long enough to judge the effect on the natural history of the disease. In 14 pregnancies, development or worsening of hypertension occurred. In nine, the diastolic blood pressure rose to 110 mm Hg or greater, and delivery was required because of hypertension. Twenty-three of 25 pregnancies ended in live births (92 percent) and 21 babies survived (84 percent). Fourteen of the 23 live births were premature. It is concluded that, in a substantial fraction of women with moderate renal insufficiency, pregnancy is accompanied by a decline in renal function; however, the fetal survival is much better than previously reported.

The role of activity blockade on glutamate receptor subunit expression in the spinal cord.

Spinal cord injury (SCI) impairs descending glutamatergic neurotransmission reaching ventral motor neurons (VMN). Previously we reported up-regulation of NMDA receptor subunits NR1 and NR2A mRNAs in VMN caudal to the lesion site 24 h after SCI. The absence of effect rostral to the injury site suggested that injury-induced loss of descending activity might be involved. To test this hypothesis, we blocked axonal conduction by focal injection of tetrodotoxin into the spinal cord. We found increased NR1 and NR2A mRNAs in VMN similar in extent to that seen after SCI. Thus, the increase in NMDA subunit mRNAs may be 'inactivity dependent' and associated with reorganization of the spinal cord in response to loss of descending innervation.

Temporal-spatial pattern of acute neuronal and glial loss after spinal cord contusion.

The secondary loss of neurons and glia over the first 24 h after spinal cord injury (SCI) contributes to the permanent functional deficits that are the unfortunate consequence of SCI. The progression of this acute secondary cell death in specific neuronal and glial populations has not previously been investigated in a quantitative manner. We used a well-characterized model of SCI to analyze the loss of ventral motoneurons (VMN) and ventral funicular astrocytes and oligodendrocytes at 15 min and 4, 8, and 24 h after an incomplete midthoracic contusion injury in the rat. We found that both the length of lesion and the length of spinal cord devoid of VMN increased in a time-dependent manner. The extent of VMN loss at specified distances rostral and caudal to the injury epicenter progressed symmetrically with time. Neuronal loss was accompanied by a loss of glial cells in ventral white matter that was significant at the epicenter by 4 h after injury. Oligodendrocyte loss followed the same temporal pattern as that of VMN while astrocyte loss was delayed. This information on the temporal-spatial pattern of cell loss can be used to investigate mechanisms involved in secondary injury of neurons and glia after SCI.

Relationship of altered glutamate receptor subunit mRNA expression to acute cell loss after spinal cord contusion.

Alterations in the expression of ionotropic glutamate receptors (GluR) contribute to neuronal loss after brain ischemia and epilepsy. In order to determine whether altered expression of GluR subunits might contribute to cell loss after spinal cord injury (SCI), we performed a time course study of subunit mRNA expression using quantitative in situ hybridization. Expression was studied in ventral horn motor neurons (VMN) and glia in adjacent ventral white matter at 15 min and 4, 8, and 24 h after SCI in tissue sections 4 mm rostral and caudal to the injury epicenter. We found that the AMPA subunit GluR2 was significantly down-regulated in VMN at 24 h, but not at the earlier times examined, although half the loss of VMN in these locations occurs by 8 h after injury. No changes in the normal expression of GluR2 or GluR4 were found in white matter where glial loss occurs after SCI. NMDA subunits NR1 and NR2A were significantly and rapidly up-regulated in VMN after SCI, but only caudal to the lesion site, while VMN loss is similar rostral and caudal to the epicenter. Thus, the temporal pattern of AMPA and the spatial pattern of NMDA subunit expression changes were distinct from the pattern of VMN loss after SCI. We conclude that altered GluR subunit expression after SCI is unlikely to be involved in secondary cell loss and instead may be involved with plasticity and reorganization of the injured spinal cord.

Expression of splice variants of the NR1 subunit of the N-methyl-D-aspartate receptor in the normal and injured rat spinal cord.

Quantitative western blot analysis in laminectomy control spinal cords of adult rats was used to provide the first report of the normal expression patterns of the N1, C1, C2 and C2' cassettes in the cervical, thoracic and lumbar regions of the spinal cord as a percent of total NR1 subunit protein. In all regions studied, the C1 and C2 cassettes were usually contained in less than 10% of total NR1 protein. In contrast, approximately 90% of total NR1 protein contained the C2' cassette. A significant proportion of total NR1 protein (approximately 30%) also contained the N1 cassette. These data are consistent with expression of NR1(000) (NR1-4a) and NR1(100) (NR1-4b) as the dominant splice forms in the spinal cord. Splice variant expression was also studied following incomplete, contusive spinal cord injury (SCI) to the thoracic level 8 (T8) region. This injury did not change expression of the C1 or C2 cassette in any region of the spinal cord acutely at 24 h or chronically at 1 month. There was an increase in expression of the N1 cassette in the lumbar regions 1 month after injury (p < 0.05). These data indicate that SCI induces distal changes in NR1 splice variant expression, which may play a role in the adaptive response of neurons in the chronically injured spinal cord.

Changes in NMDA receptor subunit expression in response to contusive spinal cord injury.

Differential assembly of N-methyl-D-aspartate (NMDA) receptor subunits determines their functional characteristics. Using in situ hybridization, we found a selective increase of the subunits NR1 and NR2A mRNA at 24 h in ventral motor neurons (VMN) caudal to a standardized spinal cord contusion injury (SCI). Other neuronal cell populations and VMN rostral to the injury site appeared unaffected. Significant up-regulation of NR2A mRNA also was seen 1 month after SCI in thoracic and lumbar VMN. The selective effects on VMN caudal to the injury site suggest that the loss of descending innervation leads to increased NMDA receptor subunit expression in these cells after SCI, which may alter their responses to glutamate. In contrast, protein levels determined by western blot analysis show decreased levels of NR2A 1 month after SCI in whole thoracic segments of spinal cord that included the injury sites. No effects of injury were seen on subunit levels in cervical or lumbar segments. Taken together with our previous study showing alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate receptor subunit down-regulation after injury, our data suggest that glutamate receptor composition is significantly altered after SCI. These changes need to be taken into account to properly understand the function of, and potential pharmacotherapy for, the chronically injured spinal cord.