Nobuo Nagano

Direct evidence for a causative role of FGF23 in the abnormal renal phosphate handling and vitamin D metabolism in rats with early-stage chronic kidney disease

Circulating levels of fibroblast growth factor 23 (FGF23) are elevated in patients with early chronic kidney disease (CKD) and are postulated to cause low blood levels of 1,25-dihydroxyvitamin D, as well as normal phosphate levels. In order to provide more direct evidence for the pathophysiological role of FGF23 in the settings of mineral ion homeostasis typically seen in early CKD, we studied rats with progressive CKD treated with anti-FGF23 neutralizing antibody. Without antibody treatment, rats with CKD exhibited high circulating levels of FGF23 and parathyroid hormone, low 1,25-dihydroxyvitamin D, and normal serum phosphate levels, accompanied by increased fractional excretion of phosphate. Antibody treatment, however, lessened fractional excretion of phosphate, thus increasing serum phosphate levels, and normalized serum 1,25-dihydroxyvitamin D by increased 1α-OHase and decreased 24-OHase expressions in the kidney. These antibody-induced changes were followed by increased serum calcium levels, leading to decreased serum parathyroid hormone. Hence, our study shows that FGF23 normalizes serum phosphate and decreases 1,25-dihydroxyvitamin D levels in early-stage CKD, and suggests a pathological sequence of events for the development of secondary hyperparathyroidism triggered by increased FGF23, followed by a reduction of 1,25-dihydroxyvitamin D and calcium levels, thereby increasing parathyroid hormone secretion.

Functional significance of subtypes of 5-HT receptors in the rat spinal reflex pathway

1. The functional significance of subtypes of 5-hydroxytryptamine (5-HT) receptors was studied in the rat spinal reflex pathway. 2. Ketanserin had no effect on the mono- (MSR) or polysynaptic reflex (PSR) in spinal rats, but decreased the PSR in intact rats. 3. 8-Hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) and 5-methoxy-N,N-dimethyltryptamine (5-MeODMT) decreased the MSR and increased the PSR in spinal rats. 4. Ketanserin antagonized the effects of 5-MeODMT without antagonizing the effects of 8-OH-DPAT. 5. Cinanserin had similar effects to those of ketanserin. 6. These results suggest that both 5-HT1A and 5-HT2 receptors mediate MSR inhibition and PSR augmentation in the spinal reflexes of spinal rats, and that the 5-HT2 receptor has a supraspinal tonic excitatory influence on the PSR in intact rats.

Sensitivity of spinal reflexes to TRH and 5-HT in 5,6-dihydroxytryptamine-treated rats

Destruction of descending serotonergic nerve terminals containing thyrotropin-releasing hormone (TRH) was affected in rats by the intracisternal injection of 5,6-dihydroxytryptamine (5,6-DHT) two weeks before subsequent experiments. Although the level of TRH in the lumbar enlargement was significantly reduced in 5,6-DHT-treated rats, the effects of TRH on the monosynaptic reflex (MSR) and the polysynaptic reflex (PSR) in these rats were no different from those in control rats. MSR inhibition by 5-methoxy-N,N-dimethyltryptamine (5-MeODMT) was attenuated by 5,6-DHT treatment although there was no obvious difference in the effects of 5-MeODMT on the PSR between 5,6-DHT-treated and control rats. In 5,6-DHT-treated rats, L-5-hydroxytryptophan (5-HTP) markedly decreased the MSR and increased the PSR although the same doses of 5-HTP did not produce any effects on either the MSR or the PSR in control rats. In control rats, after administration of imipramine or clorgyline, 5-HTP produced effects similar to those observed in 5,6-DHT-treated rats. These results suggest that the supersensitivity to 5-HTP in 5,6-DHT-treated rats is due to a lack of 5-hydroxytryptamine (5-HT) uptake into 5-HT-containing nerve terminals rather than to a change in 5-HT receptors.

The spinal reflex of chronic spinal rats is supersensitive to 5-HTP but not to TRH or 5-HT agonists.

The effects of thyrotropin-releasing hormone (TRH), 5-methoxy-N,N-dimethyltryptamine (5-MeODMT) and L-5-hydroxytryptophan (5-HTP) were studied on the monosynaptic reflex (MSR) and the polysynaptic reflex (PSR) in acute and chronic spinal rats. Radioimmunoassay showed that while chronic spinal transection (for 2 weeks) caused the complete depletion of TRH in the ventral lumbar enlargement a certain level of TRH was maintained in the dorsal lumbar enlargement. This result suggested the existence of TRH-containing neurons in the dorsal horn other than the medullary raphe neurons descending to the spinal cord. The latency to the start of the MSR was shortened in chronic spinal rats and the amplitudes of the MSR and PSR were significantly greater than those in acute spinal rats. There was no obvious difference in the effects of TRH and 5-MeODMT on spinal reflexes of acute and of chronic spinal rats although marked supersensitivity to 5-HTP was observed in both the MSR and the PSR in chronic spinal rats. The supersensitivity to 5-HTP was considered to be due to a lack of 5-hydroxytryptamine (5-HT) uptake into 5-HT-containing nerve terminals rather than to a change in 5-HT receptors. It is suggested that TRH and 5-HT do not show any mutual requirement for each other in their effects on the spinal reflex since co-depletion of TRH and 5-HT did not change the effects of TRH and 5-MeODMT in chronic spinal rats. The coexistence of 5-HT and TRH in the descending spinal pathway is not considered to be significant for the control of spinal reflexes at the postsynaptic level.

Effects of a new analog of thyrotropin-releasing hormone, tNα-[{(s)-4-oxo-2-azetidinyl} carbonyl]-l-histidyl-l-prolinamide dehydrate (YM-14673) on spinal reflex potentials and flexor reflexes in spinalized rats

Experiments were performed on spinalized rats, transected at the Cl level. The intravenous administration of TRH and its analog YM-14673 (N alpha-[(S)-4-oxo-2-azetidinyl) carbonyl]-L-histidyl-L-prolinamide dehydrate) produced marked increases in the amplitude of mono- and polysynaptic reflex potentials and those of the withdrawal flexor reflexes. The effects of YM-14673 were stronger and longer-lasting than those of TRH. The stimulant action of TRH and YM-14673 on the flexor reflexes was not antagonized by prazosin, chlorpromazine, haloperidol or cyproheptadine, suggesting no involvement of the release of catecholamines or serotonin in the stimulant effects of TRH and its analog. Therefore, YM-14673 may be beneficial for the treatment of several spinal motor neuron diseases.

Enkephalin but not morphine modulates the motor activity in the frog spinal cord in vitro

In the isolated frog spinal cord, methionine-enkephalin (ME, up to 3 X 10(-5) M) hyperpolarized the resting ventral and dorsal root potentials. These hyperpolarizations remained even under Ca2+-free conditions. ME depressed the fast component of the electrically stimulated spinal reflex and enlarged the following depolarizing component. Morphine (10(-4) M) had no apparent ME-like effects. ME may directly and indirectly affect the motoneuron and modulate the spinal motor activity in the frog spinal cord.

Different spinal effects of opioid agonists on spinal and spino-bulbo-spinal reflexes in rats

The effects of morphine-HCl (MOR), methionine-enkephalin (ME) and dynorphin1–13 (DYN) on spinal and spino-bulbo-spinal (SBS) reflexes were studied. Although spinal intrathecal administration of MOR (15μg) did not produce any apparent effect on these reflexes, systemically administered MOR (3mg/kg i.v.) reduced the electrical toe stimulation-induced SBS reflex. Furthermore, MOR (3mg/kg i.v.) increased the polysynaptic reflex induced by electrical stimulation of low-threshold dorsal root afferents in intact (non-spinal) rats, but not in spinal rats. Intrathecally administred DYN (0.5 and 5 μg) reduced both the electrical toe stimulation-induced spinal and SBS reflexes, while ME (15μg) only reduced the SBS reflex. These results indicate the physiological multiplicity of spinal opioid receptors. MOR may affect supraspinal nuclei but not the spinal pathway which possesses MOR-sensitive opioid receptors, whereas ME and DYN affect spinal opioid peptide receptors and modulate the reflex activities in which they participate.

Inhibitory effect of phencyclidine on rat spinal reflexes

1. The effects of phencyclidine (PCP) on spinal reflexes and the excitability of motoneuron somata were studied in rats. 2. PCP decreased both the monosynaptic reflex (MSR) and the polysynaptic reflex (PSR) in spinal rats as well as in intact rats. 3. The effects of PCP were not antagonized by monoaminergic and cholinergic antagonists. 4. A low dose of PCP antagonized the PSR augmentation but not the MSR augmentation caused by N-methyl-D-aspartate (NMDA). 5. In the excitability test, PCP decreased not only the monosynaptic response but also the excitability of motoneuron somata. 6. These results suggest that the inhibitory effects of PCP on spinal reflexes are not due to mediation of the monoaminergic and cholinergic systems, but partly due to its blockade of NMDA-type receptors and also to a hyperpolarizing action or a membrane-stabilizing action on the motoneuron soma.

Effects of L-threo-dihydroxyphenylserine on spinal reflexes and decerebrate rigidity in rats.

1. The effects of a noradrenaline precursor, L-threo-3,4-dihydroxyphenylserine (L-DOPS), on spinal mono-(MSR) and polysynaptic reflexes (PSR) and decerebrate rigidity, were studied. 2. Although a low dose of L-DOPS (10 mg/kg, i.v.) did not affect MSR or PSR in C1 spinal rats, high doses of L-DOPS (50 and 100 mg/kg, i.v.) moderately enhanced the amplitudes of both MSR and PSR. 3. Clorgyline-HCl (1 mg/kg, i.v.), an MAO inhibitor, enhanced the excitatory effects of low-dose L-DOPS (10 mg/kg, i.v.) on both reflexes. 4. Benserazide-HCl (1 mg/kg, i.v.), an L-aromatic amino acid decarboxylase inhibitor, decreased the pressor effect, but not the stimulatory effects, of L-DOPS (100 mg/kg, i.v.) on MSR and PSR. 5. L-DOPS (300 mg/kg, i.p. or i.d.) did not affect the muscle tone of rigid hindlimbs caused by radio frequency lesioning of the midbrain. 6. These results suggest that the moderate enhancing effects of L-DOPS on MSR and PSR are due to conversion of L-DOPS to noradrenaline in the spinal cord.