Regional Distribution of Immunoreactive-Thyrotrophin-Releasing Hormone and Substance P, and Indoleamines in Human Spinal Cord

The regional distributions of thyrotrophin-releasing hormone (TRH) and substance P in postmortem human spinal cord were determined by radioimmunoassay in fresh tissue taken from 22 patients who died without known neurological disease. Dorsal, ventral, and intermediolateral spinal cord regions were obtained from different segmental levels (lumbar L1, 2, 3, and 4; thoracic groups T1-3, T4-6, T7-9, and T10-12) together with selective regions of grey matter of lumbar spinal cord. The effects on peptide levels of the age of the patient, the postmortem time interval, and freezing the tissue samples prior to assay were assessed. Levels of 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) were determined in regional lumbar and thoracic tissue using HPLC with electrochemical detection. Substance P was found in the highest concentration in the dorsal spinal cord, with no significant segmental differences. In contrast, TRH was present in higher levels in the ventral rather than the dorsal spinal cord, with segmental differences. There was a significant difference in the 5-HT/5-HIAA ratio between dorsal and ventral spinal cord, with the highest ratio in the ventral spinal cord. There were no significant differences in substance P, TRH, or 5-HT levels in spinal cords between 5 and 20 h postmortem or from patients aged between 65 and 90 years. Freezing the tissue (-80 degrees C for 24 h) prior to assay significantly reduced TRH and substance P levels compared to samples assayed immediately without prior freezing.(ABSTRACT TRUNCATED AT 250 WORDS)     

A comparison of microdistributions of taurine and cysteine sulphinate decarboxylase activity with those of GABA and L-glutamate decarboxylase activity in rat spinal cord and thalamus

Abstract— Distribution profiles of taurine and activity of cysteine sulphinate decarboxylase (CSD), the enzyme catalysing the formations of hypotaurine from cysteine sulphinate and of taurine from cysteate respectively, in the rat spinal cord and thalamus were studied in comparison with those of GABA and activity of l -glutamate decarboxylase (GAD), the rate limiting enzyme for GABA formation. In the spinal cord (L₂-L₃), it was found that taurine is fairly evenly distributed, whereas the activity of CSD is higher in the dorsal half of the spinal cord than in the ventral half. The highest CSD activity was found in the dorsal part of the dorsal horn. In the anterior part (A 5.4) of the thalamus, taurine and CSD activity were also distributed evenly and no areas having high taurine content and CSD activity were detected. In contrast with the even distributions of taurine and CSD activity, both GABA and GAD activity were distributed unevenly in the same CNS areas examined: The areas having high GABA content and GAD activity in the thalamus (A 5.4) coincided with the ventrolateral part of the ventral nucleus of thalamus (VM), entopeduncular nucleus (EP) and nucleus reuniens thalami (RE), whereas those in the spinal cord were found to be in the dorsal part of the dorsal horn and surrounding parts of the central canal, respectively. Considering a probable role of GABA in mammalian central nervous system (CNS) as an inhibitory neurotransmitter, it seems unlikely that taurine acts as an inhibitory neurotransmitter at least in the rat spinal cord and thalamus.     

Acute administration of alcohol modulates pyroglutamyl amino peptidase II activity and mRNA levels in rat limbic regions

Released TRH is inactivated by an ectopeptidase, pyroglutamyl aminopeptidase II (PPII). PPII expression and activity are stringently regulated in adenohypophysis, and in rat brain, during kindling stimulation that activates TRHergic neurons. To gain further insight into the possible regulation of PPII, we studied the effect of an acute intraperitoneal ethanol administration that affects TRH content and expression. PPII activity was determined by a fluorometric assay and PPII mRNA levels by semi-quantitative RT-PCR. Activity decreased in frontal cortex 1 h after ethanol injection and, after 6 h, in hippocampus, amygdala and n. accumbens. PPII mRNA levels decreased at 30 and 60 min in frontal cortex and n. accumbens while increased at longer times in these regions and, in hippocampus and hypothalamus. NMDA and GABA(A) receptors' agonists and antagonists were tested at 1 h (+/-ethanol) on PPII activity and mRNA levels, as well as on TRH content and its mRNA. In n. accumbens, PPII mRNA levels decreased by ethanol, MK-801, and muscimol while picrotoxin or NMDA reversed ethanol's inhibition. Ethanol decreased TRH content and increased TRH mRNA levels as MK-801 or muscimol did (NMDA or picrotoxin reverted the effect of ethanol). In frontal cortex, PPII activity was inhibited by ethanol, NMDA and MK-801 with ethanol; its mRNA levels were reduced by ethanol, MK-801 and muscimol (NMDA and picrotoxin reverted ethanol's inhibition). These results show that PPII expression and activity can be regulated in conditions where TRHergic neurons are modulated. Effects of ethanol on PPII mRNA levels as well as those of TRH and its mRNA may involve GABA or NMDA receptors in n. accumbens. Changes observed in frontal cortex suggest combined effects with stress. The response was region-specific in magnitude, tendency and kinetics. These results give further support for brain PPII regulation in conditions that modulate the activity of TRHergic neurons.     

Purification of a specific inhibitor of pyroglutamyl aminopeptidase II from the marine annelide Hermodice carunculata. in vivo effects in rodent brain

An inhibitor of the metallo-ectoenzyme, pyroglutamyl aminopeptidase II (PPII), a thyrotropin releasing hormone-specific peptidase, was identified by screening extracts from marine species of the Cuban coast-line belonging to the phylla Chordata, Echinodermata, Annelida, Mollusca, Cnidaria, Porifera, Chlorophyta and Magnoliophyta. Isolation of the inhibitor (HcPI), from the marine annelide Hermodice carunculata, was achieved by trichloroacetic acid treatment of the aqueous extract, followed by ion-exchange chromatography on DEAE Sephacel, gel filtration on Sephadex G-25 and reverse phase-HPLC. HcPI had a small apparent molecular weight (below 1000 Da) and was not a peptide. It inhibited rat PPII (a membrane preparation with 8.5mg protein/ml) with an apparent K(i) of 51 nM. HcPI did not inhibit serine (trypsin, chymotrypsin, elastase and dipeptidyl aminopeptidase IV), cysteine (papain, bromelain and pyroglutamyl aminopeptidase I), aspartic (pepsin and recombinant human immunodeficiency virus 1 protease (HIV1-PR)) nor other metallo proteinases (collagenase, gelatinase, angiotensin converting enzyme, aminopeptidase N and carboxypeptidase A). HcPI was non-toxic and active in vivo. Intraperitoneal injection of HcPI reduced mouse pituitary and brain PPII activity. Potency of the effect was higher in hypophysis and hypothalamus than in other brain regions. Intrathecal administration to male rats reduced PPII activity in the spinal cord. In conclusion we have identified a specific inhibitor of PPII that is the first M1 family zinc metallo-peptidase inhibitor isolated from marine invertebrates. It may be useful for elucidating the in vivo role of PPII in the pituitary and central nervous system.     

Chronic ethanol or glucose consumption alter TRH content and pyroglutamyl aminopeptidase II activity in rat limbic regions

Thyrotropin-releasing hormone (TRH), its receptors and inactivating enzyme (PPII) are present in limbic regions. Nutritional changes or acute ethanol administration in male rats differentially modulate TRH or PPII expression. Chronic ethanol effect was studied in male (3, 6 and 8 weeks) and female rats (6 weeks) including naive and pair-fed (glucose) groups. Daily solid food and liquid intake, serum TSH and corticosterone, TRH content and PPII activity in limbic regions, were quantified. Gender differences were found in ethanol and total caloric intake and body weight gain, TSH and corticosterone levels. Ethanol consumption decreased TRH content and PPII activity in frontal cortex of male rats after 3-6 weeks. In contrast, glucose ingestion altered, by the third week, TRH content in amygdala, hippocampus, hypothalamus and nucleus accumbens, PPII activity in hippocampus and frontal cortex; by the sixth week, TRH content in amygdala and n. accumbens of male and females. Withdrawal at 24 h after 3-week ethanol ingestion decreased TRH content in amygdala and PPII activity in n. accumbens, while withdrawal from glucose reverted some of the effects produced by chronic glucose ingestion. Variations in TRH content or PPII activity support a region specific involvement of TRH neurons that depend on the treatment.     

Regional and Subcellular Distribution of Cyanide Metabolizing Enzymes in the Central Nervous System

Activities of cyanide metabolizing enzymes were measured in various subcellular fractions and regions in the central nervous system. Brain rhodanese and liver beta-mercaptopyruvate sulfurtransferase showed a slight decrease in activity after death. The activity of beta-mercaptopyruvate sulfurtransferase was negligible in the rat brain, compared with that of rhodanese. A small amount of thiocyanate was produced from cyanide and beta-mercaptopyruvate in the human brain, probably due to contamination with red blood cells. Rhodanese activity was widely distributed in all the areas of nervous tissue examined. In the rat the olfactory bulb showed the highest rhodanese activity, and high activity was also observed served in the thalamus, septum, hippocampus, and dorsal part of the midbrain. Rhodanese activity was low in various parts of the cerebral cortex. The distribution pattern of rhodanese in post-mortem human brain was essentially similar to that in rat brain. The thalamus, amygdala, centrum semiovale, colliculus superior, and cerebellar cortex showed high rhodanese activity in the human brain. Rhodanese activity was detected in the spinal cord. Anterior horn showed the highest rhodanese activity in the cervical, thoracic, and lumbar cord. Most rhodanese activity in the rat brain was recovered in the mitochondrial fraction with the highest specific activity. Rhodanese activity was lower in spinal cords obtained from autopsied cases with amyotrophic lateral sclerosis than in those of control subjects. A significant decrease in rhodanese was observed in the posterior column of the cervical or thoracic cord, but the activity in the anterior horn did not differ significantly between the two groups.     

Immunohistochemical investigation of neuropeptides in the central nervous system of the amphioxus,Branchiostoma belcheri

The immunohistochemical localization of nine different neuropeptides was studied in the central nervous system of the amphioxus, Branchiostoma belcheri. In the brain, perikarya immunoreactive for urotensin I and FMRFamide were localized in the vicinity of the central canal. One of the processes of each of these perikarya was found to cross the dorso ventral slit-like lumen of the central canal. Oxytocin-immunoreactive short fibers, but not perikarya, were detected in the ventral part of the brain. Perikarya immunoreactive for arginine vasopressin/vasotocin, oxytocin and FMRFamide were widely distributed in the spinal cord. Arginine vasopressin/vasotocin-immunoreactive fibers often made contacts with Rohde cell axons. Angiotensin II-immunoreactive perikarya were observed in the posterior half of the spinal cord, and urotensin I-immunoreactive perikarya were found in the caudal region of the spinal cord. Cholecystokinin/gastrin-immunoreactive fibers, but not perikarya, were detected in the spinal cord; some extended as far as the ependymal layer of the cerebral ventricle. No colocalization of the peptides examined was observed. No immunoreactivity for atrial and brain natriuretic peptides nor for urotensin II was detected. The present study indicates that there are at least six separate neuronal systems that contain different peptides, respectively, in the central nervous system of the amphioxus. Their functions remain to be determined.Part of this investigation has previously been presented in abstract form (Uemura et al. 1989)     

2',3'-Cyclic nucleotide 3'-phosphohydrolase in the developing chick brain and spinal cord

Developmental changes of the 2',3'-cyclic nucleotide 3'-phosphohydrolase activity in the chick brain and spinal cord are reported. The greater part of increase in enzyme activity occurred between 18 days of incubation and 3 days after hatching in the whole brain, and between 18 and 21 days of incubation in the spinal cord. These periods are those of active myelination in the chick brain and spinal cord, respectively. The possibility was emphasized that 2',3'-cyclic nucleotide 3'-phosphohydrolase can be used as a marker for the myelin sheaths in the developing central nervous system. Comparisons were also made among the developmental changes in the forebrain, midbrain, brain stem, cerebellum, and spinal cord.     

Amygdala kindling differentially regulates the expression of the elements involved in TRH transmission

Subthreshold electrical stimulation of the amygdala (kindling) activates neuronal pathways increasing the expression of several neuropeptides including thyrotropin releasing-hormone (TRH). Partial kindling enhances TRH expression and the activity or its inactivating ectoenzyme; once kindling is established (stage V), TRH and its mRNA levels are further increased but TRH-binding and pyroglutamyl aminopeptidase II (PPII) activity decreased in epileptogenic areas. To determine whether variations in TRH receptor binding or PPII activity are due to regulation of their synthesis, mRNA levels of TRH receptors (R1, R2) and PPII were semi-quantified by RT-PCR in amygdala, frontal cortex and hippocampus of kindled rats sacrificed at stage II or V. Increased mRNA levels of PPII were found at stage II in amygdala and frontal cortex, and of pro-TRH and TRH-R2, in amygdala and hippocampus. At stage V, pro-TRH mRNA levels increased and those of PPII, decreased in the three regions; TRH-R2 mRNA levels diminished in amygdala and frontal cortex and of TRH-R1 only in amygdala. In situ hybridization analyses revealed, at stage II, enhanced TRH-R1 mRNA levels in dentate gyrus and amygdala while decreased in piriform cortex; those of TRH-R2 increased in amygdala, CA2, dentate gyrus, piriform cortex, thalamus and subiculum and of PPII, in CAs and piriform cortex. In contrast, at stage V decreased expression of TRH-R1 occurred in amygdala, CA2/3, dentate gyrus and piriform cortex; of TRH-R2 in CA2, thalamus and piriform cortex, and of PPII in CA2, and amygdala. The magnitude of changes differed between ipsi and contralateral side. These results support a trans-synaptic modulation of all elements involved in TRH transmission in conditions that stimulate the activity of TRHergic neurons. They show that reported changes in PPII activity or TRH-binding caused by kindling relate to regulation of the expression of TRH receptors and degrading enzyme.     

3,3',5'-triiodo-L-thyronine reduces efficiency of mRNA knockdown by antisense oligodeoxynucleotides: a study with pyroglutamyl aminopeptidase II in adenohypophysis

The impact of hormones on the efficacy of antisense oligodeoxynucleotides (ASOs) is a poorly analyzed subject. We designed, based on the identification of potentially favorable local elements of mRNA secondary structure, eight phosphorothioate ASOs to knock down the expression of an ectopeptidase, pyroglutamyl aminopeptidase II (PPII), in primary cultures of adenohypophysis. Two of the PPII ASOs were very efficient, sequence-specific, and target-specific. Because the expression of PPII is upregulated by 3,3',5'-triiodo-L-thyronine (T3), we studied the impact of varying the protocol of PPII induction on the knockdown efficacy. Hormone removal at transfection increased markedly the ability of (1) PPII ASOs to reduce PPII mRNA levels or PPII activity in adenohypophyseal cells or in C6 rat glioma cells and (2) a thyrotropin-releasing hormone (TRH) receptor-1 (TRH-R1) ASO to reduce TRH-R1 mRNA levels in adenohypophyseal cells. There was no effect of hormone removal on transfection efficacy and no correlation between target mRNA levels and ASO efficacy. These data demonstrated that ASO efficacy could depend on T3 levels; this might be due to regulation of a step generally critical for ASO efficiency.     

Encephalomyelitis in mice experimentally infected with Akabane virus.

Lesions in the central nervous system of mice, induced by intracerebral injection of Akabane virus, were observed by the fluorescent antibody technique and histological method. Fluorescent antigens were recognized in the cytoplasm of nerve cells, but were not detected exactly in any other part. Fluoresced nerve cells were distributed almost all over the central nervous system, especially in medulla oblongata and spinal cord. The appearance of fluorescent antigens was followed by histological changes. So-called Nissl's acute severe degeneration was observed in nerve cells in the area where the fluorescent antigens were distributed. Spongy foci were seen in medulla oblongata and spinal cord. Virus was recovered from brain and spinal cord, but not from any other visceral organ or blood. Akabane virus showed an affinity to nerve cells and caused primary nonpurulent encephalomyelitis when inoculated intracerebrally to mice.     

Biochemical and autoradiographic studies of TRH receptors in sections of rabbit spinal cord

Receptors for thyrotropin-releasing hormone (pGlu-His-Pro-NH2, TRH) on thaw-mounted sections of rabbit spinal cord have been identified biochemically and visualized by light microscopic autoradiography. Binding of [3H] [3-Me-His2]TRH to 20 microns sections exhibited high apparent affinity and a pharmacological specificity almost identical to that previously demonstrated for spinal TRH receptors in membranes. In autoradiograms, the highest density of TRH receptors appeared in the substantia gelatinosa of the dorsal gray and around the central canal, with intermediate levels in the ventral gray.     

Alendronate lowers cholesterol synthesis in the central nervous system of rats - a preliminary study

Nitrogen-containing bisphosphonates were found to inhibit farnesyl diphosphate synthase - an essential enzyme in the cholesterol biosynthesis pathway, but their effect on cholesterol synthesis per se in the central nervous system (CNS) remains unknown. The aim of the present study was to examine possible influence of a representative agent alendronate on cholesterol synthesis rates in selected parts of rat CNS and on plasma cholesterol level. Two groups of rats were orally administered either alendronate (3 mg/kg b.w.) or vehicle for 9 days. At the end of experiment, brain (basal ganglia, frontal cortex and hippocampus) and spinal cord were isolated and cholesterol synthesis was determined using the technique of deuterium incorporation from deuterated water. In the alendronate group significant reductions of cholesterol synthesis rates were detected in frontal cortex, hippocampus and spinal cord (p<0.001). However, the experimental treatment did not produce a significant alteration in the levels of plasma cholesterol. In conclusion, this study brings the first experimental evidence of the inhibition of cholesterol biosynthesis with alendronate in central nervous system.     

Retrograde axonal transport of an exogenous enzyme covalently linked to B-IIb fragment of tetanus toxin.

Attempt to replace enzymes in a number of fatal lysosomal storage disease involving the central nervous system have as yet been unsuccessful owing to the impermeability of the blood/brain barrier to macromolecules. In order to treat storage disease due to enzyme deficiencies, we investigated the feasibility of transporting an enzyme into the central nervous system without crossing the blood/brain barrier. Using the B-IIb fragment of tetanus toxin (because it is involved in recognition by the nerve-cell endings), retrograde axonal transport toward the spinal cord and trans-synaptic movement, and glucose oxidase as a marker, we demonstrated that a non-toxic enzyme-vector conjugate was taken up by axon terminals. After injection into the gastrocnemius muscle, the B-IIb-glucose oxidase conjugate was detected, both histologically and electrochemically, distally to a ligature on the sciatic nerve. Thus the B-IIb fragment could serve as a vector for glucose oxidase transport into the central nervous system. It was also verified that the transported enzyme retained its activity. Transport of this 150 kDa molecule by fragment B-IIb of tetanus toxin suggests that other enzymes of a lesser molecular mass may also be transported.     

Susceptibility of chickens to avian encephalomyelitis virus. IV. Behavior of the virus in laying hens.

Some laying hens 6 months of age were inoculated subcutaneously or orally with a chick embryo--adapted strain of avian encephalomyelitis virus and examined for propagation of the virus in the body. When inoculated subcutaneously, the virus appeared in liver, spleen, ovarian follicle, and muscle at the site of inoculation 1 day, in kidney and lumbar part of the spinal cord 3 days, in the pancreas 5 days, in heart, duodenum, and cervical part of the spinal cord 7 days, and in the brain 11 days after inoculation. After its appearance, it increased gradually in amount in liver, spleen, pancreas, muscle at the site of inoculation, and cervical and lumbar parts of the spinal cord, but remained at a low level in any other organ. When examined 14 days after inoculation and later, it was distributed mainly in the central nervous system. It was detected from 12 of 16 organs examined. The highest virus level in each organ was 10(2.6)/0.1 g in pancreas and lumbar part of the spinal cord, which were followed by muscle at the site of inoculation (10(2.0)/0.1 g), spleen (10(1.8)/0.1 g), cervical part of the spinal cord, heart, and liver in the order listed. When inoculated orally, the virus was found sporadically in spleen, pancreas, kidney, cecum, ovarian follicle, and lumbar part of the spinal cord. The virus level was low in these organs, of which pancreas, kidney, and lumbar part of the spinal cord showed the highest virus level, or 10(1.3)/0.1 g.     

Brain and spinal cord interaction: protective effects of exercise prior to spinal cord injury

We have investigated the effects of a spinal cord injury on the brain and spinal cord, and whether exercise provided before the injury could organize a protective reaction across the neuroaxis. Animals were exposed to 21 days of voluntary exercise, followed by a full spinal transection (T7-T9) and sacrificed two days later. Here we show that the effects of spinal cord injury go beyond the spinal cord itself and influence the molecular substrates of synaptic plasticity and learning in the brain. The injury reduced BDNF levels in the hippocampus in conjunction with the activated forms of p-synapsin I, p-CREB and p-CaMK II, while exercise prior to injury prevented these reductions. Similar effects of the injury were observed in the lumbar enlargement region of the spinal cord, where exercise prevented the reductions in BDNF, and p-CREB. Furthermore, the response of the hippocampus to the spinal lesion appeared to be coordinated to that of the spinal cord, as evidenced by corresponding injury-related changes in BDNF levels in the brain and spinal cord. These results provide an indication for the increased vulnerability of brain centers after spinal cord injury. These findings also imply that the level of chronic activity prior to a spinal cord injury could determine the level of sensory-motor and cognitive recovery following the injury. In particular, exercise prior to the injury onset appears to foster protective mechanisms in the brain and spinal cord.     

Molecular characterization of a novel type of prostamide/prostaglandin F synthase, belonging to the thioredoxin-like superfamily

Prostaglandin F (PGF) ethanolamide (prostamide F) synthase, which catalyzed the reduction of prostamide H(2) to prostamide F(2alpha), was found in mouse and swine brain. The enzyme was purified from swine brain, and its amino acid sequence was defined. The mouse enzyme consisted of a 603-bp open reading frame coding for a 201-amino acid polypeptide with a molecular weight of 21,669. The amino acid sequence placed the enzyme in the thioredoxin-like superfamily with Cys(44) being the active site. The enzyme expressed in Escherichia coli as well as the native enzyme catalyzed not only the reduction of prostamide H(2) to prostamide F(2alpha) but also that of PGH(2) to PGF(2alpha). The V(max) and K(m) values for prostamide H(2) were about 0.25 micromol/min.mg of protein and 7.6 microm, respectively, and those for PGH(2) were about 0.69 micromol/min.mg of protein and 6.9 microm, respectively. Neither PGE(2) nor PGD(2) served as a substrate for this synthase. Based on these data, we named the enzyme prostamide/PGF synthase. Although the enzyme showed a broad specificity for reductants, reduced thioredoxin preferentially served as a reducing equivalent donor for this enzyme. Moreover, Northern and Western blot analyses in addition to the prostamide F synthase activity showed that the enzyme was mainly distributed in the brain and spinal cord, and the immunohistochemical study in the spinal cord showed that the enzyme was found mainly in the cytosol. These results suggest that prostamide/PGF synthase may play an important functional role in the central nervous system.     

Extrahypothalamic thyrotropin releasing hormone (TRH) -- its distribution and its functions.

Thyrotropin releasing hormone (TRH) is distributed throughout the extrahypothalamic nervous system and spinal cord, in the retina, in the pancreas and gastrointestinal tract, in the placenta, in amniotic fluid, in the adrenals and in frog skin. TRH has been shown to have a variety of effects in the central nervous system, both on isolated neurones and in a number of in vivo situations. TRH interacts with endogenous and exogenous opiates and it has been suggested that endogenous TRH may mediate part of the opiate withdrawal syndrome. The presence of TRH in the retina suggests the possibility that TRH plays a role in the visual process. TRH appears to be integrally related to central thermoregulatory mechanisms. The role of TRH in psychiatric disorders is at present controversial. Recent studies suggest a role for TRH as a modulator of gastrointestinal and pancreatic function. The gastrointestinal actions of TRH include inhibition of gastric acid secretion and alterations in gastic motility. The high concentrations of TRH in the neonatal pancreas suggest a role for TRH in the early development of the pancreas. One of the metabolites of TRH histidyl-proline diketopiperazone, appears to have a number of extrahypothalamic actions and this suggests the need for further exploration of the affects of this compound both on the central nervous system and the gastrointestinal tract. The multiple extrahypothalamic actions of TRH have led to the concept that it is an ubiquitous neurotransmitter that has been co-opted by the pituitary as a releasing factor.     

Glutamic Acid Decarboxylase in Sea Lamprey (Petromyzon marinus): Characterization, Localization, and Developmental Changes

Abstract: We have carried out assays for glutamic acid decarboxylase (GAD) in homogenates of brain and spinal cord from larval and adult sea lamprey ( Petromyzon marinus ). The enzyme had similar characteristics in both stages. Optimal pH was 6.8; optimal temperature was 27–30° C; K _m at 27°C was 5 mM. GAD activity was distributed uniformly along the length of the spinal cord. Specific activities for the larval cord and brain were 26 and 63 nm CO₂/mg protein/h. respectively. The specific activities for the adult cord and brain were 29 and 236 nm CO₂/mg protein/h, respectively. Thus, the activity of cord homogenates did not change significantly between larval and adult stages, but that of the brain increased about fourfold.