A Veeranna, Ph.D.

Perinatal abrogation of Cdk5 expression in brain results in neuronal migration defects
Hirasawa M; Ohshima T; Takahashi S; Longenecker G; Honjo Y; Veeranna; Pant HC; Mikoshiba K; Brady RO; Kulkarni AB
2004 Apr 20;101(16):6249-6254, Proceedings of the National Academy of Sciences of the United States of America
Cyclin-dependent kinase 5 (Cdk5) is essential for the proper development of the CNS, as is evident from the perinatal lethality of conventional Cdk5 knockout (Cdk5-/-) mice. Cdk5 is also implicated in numerous complex functions of the adult CNS such as synaptic transmission, synaptic plasticity, and neuronal signaling. To elucidate the molecular roles of Cdk5 in the adult CNS, we have abrogated neuronal expression of Cdk5 in perinatal mice by using a cre-loxP system. The Cdk5-loxP flanked mice were crossed with the cre-transgenic mice in which the cre expression is driven by the murine neurofilament-heavy chain promoter, resulting in generation of viable Cdk5 conditional knockout mice with the restricted deletion of the Cdk5 gene in specific neurons beginning around embryonic day 16.5. Twenty-five percent of the Cdk5 conditional knockout mice carrying the heterozygous cre allele had neuronal migration defects confined to brain areas where neuronal migration continues through the perinatal period. These results indicate that abrogation of Cdk5 expression in mature neurons results in a viable mouse model that offers further opportunities to investigate the molecular roles of Cdk5 in the adult CNS
— id: 61274, year: 2004, vol: 101, page: 6249, stat: Journal Article,

Calpain mediates calcium-induced activation of the erk1,2 MAPK pathway and cytoskeletal phosphorylation in neurons: relevance to Alzheimer's disease
Veeranna; Kaji T; Boland B; Odrljin T; Mohan P; Basavarajappa BS; Peterhoff C; Cataldo A; Rudnicki A; Amin N; Li BS; Pant HC; Hungund BL; Arancio O; Nixon RA
2004 Oct;165(3):795-805, American journal of pathology
Aberrant phosphorylation of the neuronal cytoskeleton is an early pathological event in Alzheimer's disease (AD), but the underlying mechanisms are unclear. Here, we demonstrate in the brains of AD patients that neurofilament hyperphosphorylation in neocortical pyramidal neurons is accompanied by activation of both Erk1,2 and calpain. Using immunochemistry, Western blot analysis, and kinase activity measurements, we show in primary hippocampal and cerebellar granule (CG) neurons that calcium influx activates calpain and Erk1,2 and increases neurofilament phosphorylation on carboxy terminal polypeptide sites known to be modulated by Erk1,2 and to be altered in AD. Blocking Erk1,2 activity either with antisense oligonucleotides to Erk1,2 mRNA sequences or by specifically inhibiting its upstream activating kinase MEK1,2 markedly reduced neurofilament phosphorylation. Calpeptin, a cell-permeable calpain inhibitor, blocked both Erk1,2 activation and neurofilament hyperphosphorylation at concentrations that inhibit calpain-mediated cleavage of brain spectrin. By contrast, inhibiting Erk1,2 with U-0126, a specific inhibitor of Mek1,2, had no appreciable effect on ionomycin-induced calpain activation. These findings demonstrate that, under conditions of calcium injury in neurons, calpains are upstream activators of Erk1,2 signaling and are likely to mediate in part the hyperphosphorylation of neurofilaments and tau seen at early stages of AD as well as the neuron survival-related functions of the MAP kinase pathway
— id: 46128, year: 2004, vol: 165, page: 795, stat: Journal Article,

Calpain mediates calcium-induced activation of the ERK 1,2 MAPK pathway and cytoskeletal phosphorylation in neurons: Relevance to Alzheimer's disease
Veeranna; Kaji, T; Boland, B; Odrljin, T; Mohan, P; Basavarajappa, BS; Peterhoff, C; Cataldo, AM; Rudnicki, A; Li, BS; Pant, HC; Hungund, BL; Arancio, O; Nixon, RA
2004 JUL ;25(10):S173-S173, Neurobiology of aging
— id: 47723, year: 2004, vol: 25, page: S173, stat: Journal Article,

Cdk5 is a key factor in tau aggregation and tangle formation in vivo
Noble W; Olm V; Takata K; Casey E; Mary O; Meyerson J; Gaynor K; LaFrancois J; Wang L; Kondo T; Davies P; Burns M; Veeranna; Nixon R; Dickson D; Matsuoka Y; Ahlijanian M; Lau LF; Duff K
2003 May 22;38(4):555-565, Neuron
Tau aggregation is a common feature of neurodegenerative diseases such as Alzheimer's disease, and hyperphosphorylation of tau has been implicated as a fundamental pathogenic mechanism in this process. To examine the impact of cdk5 in tau aggregation and tangle formation, we crossed transgenic mice overexpressing the cdk5 activator p25, with transgenic mice overexpressing mutant (P301L) human tau. Tau was hyperphosphorylated at several sites in the double transgenics, and there was a highly significant accumulation of aggregated tau in brainstem and cortex. This was accompanied by increased numbers of silver-stained neurofibrillary tangles (NFTs). Insoluble tau was also associated with active GSK. Thus, cdk5 can initiate a major impact on tau pathology progression that probably involves several kinases. Kinase inhibitors may thus be beneficial therapeutically
— id: 60993, year: 2003, vol: 38, page: 555, stat: Journal Article,

Phosphorylation of the head domain of neurofilament protein (NF-M): a factor regulating topographic phosphorylation of NF-M tail domain KSP sites in neurons
Zheng YL; Li BS; Veeranna; Pant HC
2003 Jun 27;278(26):24026-24032, Journal of biological chemistry
In neurons the phosphorylation of neurofilament (NF) proteins NF-M and NF-H is topographically regulated. Although kinases and NF subunits are synthesized in cell bodies, extensive phosphorylation of the KSP repeats in tail domains of NF-M and NF-H occurs primarily in axons. The nature of this regulation, however, is not understood. As obligate heteropolymers, NF assembly requires interactions between the core NF-L with NF-M or NF-H subunits, a process inhibited by NF head domain phosphorylation. Phosphorylation of head domains at protein kinase A (PKA)-specific sites seems to occur transiently in cell bodies after NF subunit synthesis. We have proposed that transient phosphorylation of head domains prevents NF assembly in the soma and inhibits tail domain phosphorylation; i.e. assembly and KSP phosphorylation in axons depends on prior dephosphorylation of head domain sites. Deregulation of this process leads to pathological accumulations of phosphorylated NFs in the soma as seen in some neurodegenerative disorders. To test this hypothesis, we studied the effect of PKA phosphorylation of the NF-M head domain on phosphorylation of tail domain KSP sites. In rat cortical neurons we showed that head domain phosphorylation of endogenous NF-M by forskolin-activated PKA inhibits NF-M tail domain phosphorylation. To demonstrate the site specificity of PKA phosphorylation and its effect on tail domain phosphorylation, we transfected NIH3T3 cells with NF-M mutated at PKA-specific head domain serine residues. Epidermal growth factor stimulation of cells with mutant NF-M in the presence of forskolin exhibited no inhibition of NF-tail domain phosphorylation compared with the wild type NF-M-transfected cells. This is consistent with our hypothesis that transient phosphorylation of NF-M head domains inhibits tail domain phosphorylation and suggests this as one of several mechanisms underlying topographic regulation
— id: 61275, year: 2003, vol: 278, page: 24026, stat: Journal Article,

Phosphorylation of MEK1 by cdk5/p35 down-regulates the mitogen-activated protein kinase pathway
Sharma P; Veeranna; Sharma M; Amin ND; Sihag RK; Grant P; Ahn N; Kulkarni AB; Pant HC
2002 Jan 4;277(1):528-534, Journal of biological chemistry
Cyclin-dependent protein kinase 5 (cdk5), a member of the cdk family, is active mainly in postmitotic cells and plays important roles in neuronal development and migration, neurite outgrowth, and synaptic transmission. In this study we investigated the relationship between cdk5 activity and regulation of the mitogen-activated protein (MAP) kinase pathway. We report that cdk5 phosphorylates the MAP kinase kinase-1 (MEK1) in vivo as well as the Ras-activated MEK1 in vitro. The phosphorylation of MEK1 by cdk5 resulted in inhibition of MEK1 catalytic activity and the phosphorylation of extracellular signal-regulated kinase (ERK) 1/2. In p35 (cdk5 activator) -/- mice, which lack appreciable cdk5 activity, we observed an increase in the phosphorylation of NF-M subunit of neurofilament proteins that correlated with an up-regulation of MEK1 and ERK1/2 activity. The activity of a constitutively active MEK1 with threonine 286 mutated to alanine (within a TPXK cdk5 phosphorylation motif in the proline-rich domain) was not affected by cdk5 phosphorylation, suggesting that Thr286 might be the cdk5/p35 phosphorylation-dependent regulatory site. These findings support the hypothesis that cdk5 and the MAP kinase pathway cross-talk in the regulation of neuronal functions. Moreover, these data and the recent studies of Harada et al. (Harada, T., Morooka, T., Ogawa, S., and Nishida, E. (2001) Nat. Cell Biol. 3, 453-459) have prompted us to propose a model for feedback down-regulation of the MAP kinase signal cascade by cdk5 inactivation of MEK1
— id: 48163, year: 2002, vol: 277, page: 528, stat: Journal Article,

Synergistic contributions of cyclin-dependant kinase 5/p35 and Reelin/Dab1 to the positioning of cortical neurons in the developing mouse brain
Ohshima T; Ogawa M; Veeranna; Hirasawa M; Longenecker G; Ishiguro K; Pant HC; Brady RO; Kulkarni AB; Mikoshiba K
2001 Feb 27;98(5):2764-2769, Proceedings of the National Academy of Sciences of the United States of America
Cyclin-dependent kinase (Cdk) 5 is a unique member of the Cdk family, because Cdk5 kinase activity is detected only in the nervous tissue. Two neuron-specific activating subunits of Cdk5, p35 and p39, have been identified. Overlapping expression pattern of these isoforms in the embryonic mouse brain and the significant residual Cdk5 kinase activity in brain homogenate of the p35-/- mice indicate the redundant functions of the Cdk5 activators in vivo. Severe neuronal migration defects in p35-/-Cdk5 +/- mice further support the idea that the redundant expression of the Cdk5 activators may cause a milder phenotype in p35-/- mice compared with Cdk5-/- mice. Mutant mice lacking either Cdk5 or p35 exhibit certain similarities with Reelin/Dab1-mutant mice in the disorganization of cortical laminar structure in the brain. To elucidate the relationship between Cdk5/p35 and Reelin/Dab1 signaling, we generated mouse lines that have combined defects of these genes. The addition of heterozygosity of either Dab1 or Reelin mutation to p35-/- causes the extensive migration defects of cortical neurons in the cerebellum. In the double-null mice of p35 and either Dab1 or Reelin, additional migration defects occur in the Purkinje cells in the cerebellum and in the pyramidal neurons in the hippocampus. These additional defects in neuronal migration in mice lacking both Cdk5/p35 and Reelin/Dab1 indicate that Cdk5/p35 may contribute synergistically to the positioning of the cortical neurons in the developing mouse brain
— id: 24252, year: 2001, vol: 98, page: 2764, stat: Journal Article,

Neuronal cyclin-dependent kinase 5 activity is critical for survival
Tanaka T; Veeranna; Ohshima T; Rajan P; Amin ND; Cho A; Sreenath T; Pant HC; Brady RO; Kulkarni AB
2001 Jan 15;21(2):550-558, Journal of neuroscience
Cyclin-dependent kinase 5 (Cdk5) null mice exhibit a unique phenotype characterized by perinatal mortality, disrupted cerebral cortical layering attributable to abnormal neuronal migration, lack of cerebellar foliation, and chromatolytic changes of neurons in the brainstem and the spinal cord. Because Cdk5 is expressed in both neurons and astrocytes, it has been unclear whether this phenotype is primarily attributable to defects in neurons or in astrocytes. Herein we report reconstitution of Cdk5 expression in neurons in Cdk5 null mice and its effect on the null phenotype. Unlike the Cdk5 null mice, the reconstituted Cdk5 null mice that express the Cdk5 transgene under the p35 promoter (TgKO mice) were viable and fertile. Because Cdk5 expression is mainly limited to neurons in these mice and rescues the defects in the nervous system of the Cdk5 null phenotype, it clearly demonstrates that Cdk5 activity is necessary for normal development and survival of p35-expressing neurons
— id: 24253, year: 2001, vol: 21, page: 550, stat: Journal Article,

Regulation of axonal neurofilament phosphorylation
Pant HC; Veeranna; Grant P
2000 ;36(2):133-150, Current topics in cellular regulation
— id: 24254, year: 2000, vol: 36, page: 133, stat: Journal Article,

Cdk5 and MAPK are associated with complexes of cytoskeletal proteins in rat brain
Veeranna; Shetty KT; Takahashi M; Grant P; Pant HC
2000 Mar 29;76(2):229-236, Brain research. Molecular brain research
Neurofilament proteins, the major cytoskeletal components of large myelinated axons, are highly phosphorylated by second messenger-dependent and -independent kinases. These kinases, together with tubulins and other cytoskeletal proteins, have been shown to bind to neurofilament preparations. Cdk5 and Erk2, proline-directed kinases in neuronal tissues, phosphorylate the Lys-Ser-Pro (KSP) repeats in tail domains of NF-H, NF-M, and other axonal proteins such as tau and synapsin. In neurofilament and microtubule preparations from rat brain, we demonstrated by Western blot analysis that cdk5, a neuronal cyclin dependent kinase and Erk1/2 were associated with complexes of NF proteins, tubulins and tau. Using P13(suc1) affinity chromatography, a procedure known to bind cdc2-like kinases in proliferating cells with high affinity, we obtained a P13 complex from a rat brain extract exhibiting the same profiles of cdk5 and Erk2 bound to cytoskeletal proteins. The phosphorylation activities of these preparations and the effect of the cdk5 inhibitor, butyrolactone, were consistent with the presence of active kinases. Finally, during a column fractionation and purification of Erk kinases from rat brain extracts, fractions enriched in Erk kinase activity also exhibited co-elution of phosphorylated NF-H, tubulin, tau and cdk5. We suggest that in mammalian brain, different kinases, their regulators and phosphatases form multimeric complexes with cytoskeletal proteins and regulate multisite phosphorylation from synthesis in the cell body to transport and assembly in the axon
— id: 24255, year: 2000, vol: 76, page: 229, stat: Journal Article,

Calcium influx and membrane depolarization induce phosphorylation of neurofilament (NF-M) KSP repeats in PC12 cells
Li BS; Veeranna; Grant P; Pant HC
1999 Jun 18;70(1):84-91, Brain research. Molecular brain research
Signals activating the kinases that phosphorylate neurofilament proteins in the axon remain unknown. In a previous study, we have demonstrated that a constitutively active form of MEK1 activates Erk1 and Erk2 kinases, which phosphorylate co-transfected NF-M in NIH 3T3 cells. In this study, we report the activation of endogenous Erk1 and Erk2 by membrane depolarization and calcium influx through L-type calcium channels, which resulted in phosphorylation of the NF-M tail domain in PC12 cells. This phosphorylation was inhibited in the presence of nifedipine, an L-type calcium channel inhibitor, and PD98059, a specific MEK1 inhibitor. Our data suggest a mechanism linking calcium influx through voltage-gated calcium channels with the MAP kinase pathway and NF-M tail domain phosphorylation in cell body and neurite. These findings may provide significant new insights into mechanisms involved in some neurological diseases
— id: 24256, year: 1999, vol: 70, page: 84, stat: Journal Article,

Activation of mitogen-activated protein kinases (Erk1 and Erk2) cascade results in phosphorylation of NF-M tail domains in transfected NIH 3T3 cells
Li BS; Veeranna; Gu J; Grant P; Pant HC
1999 May;262(1):211-217, European journal of biochemistry
Neurofilaments (NFs) are neuron-specific intermediate filaments, and are the major cytoskeletal component in large myelinated axons. Lysine-serine-proline (KSP) repeats in the tail domains of high molecular weight NF proteins (NF-M and NF-H) are extensively phosphorylated in vivo in the axon. This phosphorylation in the tail domain has been postulated to play an important role in mediating neuron-specific properties, including axonal caliber and conduction velocity. Recent studies have shown that the mitogen-activated protein kinases (extracellular signal-regulated kinases, Erk1 and Erk2) phosphorylate KSP motifs in peptide substrates derived from the NF-M and NF-H tail domains in vitro. However, it is not clear whether activation of the mitogen activated protein (MAP) kinase pathway is able to phosphorylate these domains in vivo. To answer this question, a constitutively active form of mitogen-activated Erk activating kinase (MEK1) was cotransfected with an NF-M expression construct into NIH 3T3 cells. The activated mutant, but not the dominant negative mutant, induced phosphorylation of NF-M. In addition, it was shown that epidermal growth factor, which induces the MAP kinase cascade in NIH 3T3 cells, also activated endogenous Erk1 and Erk2 and NF-M tail domain phosphorylation in the transfected cells. These results present direct evidence that in-vivo activation of Erk1 and Erk 2 is sufficient for NF-M tail domain phosphorylation in transfected cells
— id: 24257, year: 1999, vol: 262, page: 211, stat: Journal Article,

Phosphorylation Activity in the Alzheimer's Disease and Normal Brain is Modulated by Microtubule-Associated Protein, Tau In Vitro
Pant MK; Veeranna; Amin ND; Amin N; Pant HC
1999 ;1:169-182, Journal of Alzheimer's Disease
One of the hallmarks of Alzheimer's disease is the presence of abundant neurofibrillary tangles (NFTs) in the brains of affected individuals. Hyperphosphorylated tau is a major component of paired helical filaments (PHFs) in NFTs. Tau is a neuronal microtubule associated protein found primarily in axons. Normal tau promotes tubulin polymerization and stabilizes microtubule (MT) structures, whereas hyperphosphorylated tau reduces its affinity for MTs and destabilizes MT-structures. This results in the disruption of vital cellular processes (e.g., axonal transport) and leads to the degeneration of affected neurons. Processes leading to the hyperphosphorylation of tau and formation of neurofibrillary lesions in Alzheimer's disease (AD) brains are not understood. Phosphorylation of a substrate molecule like tau depends upon the equilibrium between kinase and phosphatase activities and the availability of their substrate molecules in a given system. Therefore, to understand the relative roles of kinase and phosphatase activities, we studied the long-term kinetics of phosphorylation in AD and control brain extracts in the presence and absence of the phosphatase inhibitor okadaic acid (OA) using histone, casein and bacterially expressed tau as exogenous substrates. It was found that both kinase and phosphatase activities were higher in AD compared to control brains. Surprisingly, between 18 and 24 hours, there was a robust increase in phosphorylation of endogenous proteins in the brain extracts only when bacterially expressed tau was present in the phosphorylation reaction mixture. This pattern of phosphorylation activity was unaffected by OA. Significant differences in the phosphorylation of tau isoforms were also seen during this period. These data suggest that the expression and differential phosphorylation of certain tau isoforms may be responsible for the robust increase in phosphorylation and may plan an important role in Alzheimer's pathology.
— id: 24774, year: 1999, vol: 1, page: 169, stat: Journal Article,

Characterization of serine and threonine phosphorylation sites in beta-elimination/ethanethiol addition-modified proteins by electrospray tandem mass spectrometry and database searching
Jaffe H; Veeranna; Pant HC
1998 Nov 17;37(46):16211-16224, Biochemistry
A new method for the characterization of serine and threonine phosphorylation sites in proteins has been developed. After modification of a phosphoprotein by beta-elimination/ethanethiol addition and conversion of phosphoserine and phosphothreonine residues to S-ethylcysteinyl or beta-methyl-S-ethylcysteinyl residues, the modified protein was subjected to proteolytic digestion. Resulting digests were analyzed by a combination of microbore liquid chromatography, electrospray ionization tandem (MS/MS) ion trap mass spectrometry and database searching to identify original phosphorylated residues. The computer program utilized (SEQUEST) is capable of identifying peptides and modified residues from uninterpreted MS/MS spectra, and using this method, all of the five known phosphorylation sites in bovine beta-casein were identified. Application of the method to multiply phosphorylated human high molecular weight neurofilament protein (NF-H) resulted in the identification of 21 peptides and their modified residues and hence, the in vivo phosphorylation sites. These included 26 KSP and 1 KTP site, all of which occur in the KSP repeat C-terminal tail domain (residues 502-823). One site at residue 518 was previously uncharacterized. A novel non-KSP serine at residue 421 near the KLLEGEE region in a IPFSLPE motif was characterized as phosphorylated (or glycosylated). The 27 characterized phosphorylation sites occur at S/TP residues in the following motifs: KSPVKEE, KSPAEAK, KSPEKEE, KSPAEVK, KSPEKAK, KSPPEAK, KSPVKAE, and KTPAKEE. On the basis of kinase consensus sequences, all of these motifs, including the previously unreported KTPAKEE motif, can be phosphorylated by proline-directed kinases. Advantages of the new method vis-a-vis our previously reported method [Jaffe, H., Veeranna, Shetty, K. T., and Pant, H. C. (1998) Biochemistry 37, 3931-3940] include (i) production of diastereomers eluting at different retention times increased the chances of peptide identification, (ii) increased hydrophobicity and hence retention time of the modified peptides, (iii) facilitation of positive ion production, and (iv) increased susceptibility to tryptic digestion as a result of conversion of negatively charged phosphorylated residues to neutral S-ethylcysteine or beta-methyl-S-ethylcysteine residues
— id: 24258, year: 1998, vol: 37, page: 16211, stat: Journal Article,

Characterization of the phosphorylation sites of human high molecular weight neurofilament protein by electrospray ionization tandem mass spectrometry and database searching
Jaffe H; Veeranna; Shetty KT; Pant HC
1998 Mar 17;37(11):3931-3940, Biochemistry
Hyperphosphorylated high molecular weight neurofilament protein (NF-H) exhibits extensive phosphorylation on lysine-serine-proline (KSP) repeats in the C-terminal domain of the molecule. Specific phosphorylation sites in human NF-H were identified by proteolytic digestion and analysis of the resulting digests by a combination of microbore liquid chromatography, electrospray ionization tandem (MS/MS) ion trap mass spectrometry, and database searching. The computer programs utilized (PEPSEARCH and SEQUEST) are capable of identifying peptides and phosphorylation sites from uninterpreted MS/MS spectra, and by use of these methods, 27 phosphopeptides and their phosphorylated residues were identified. On the basis of these phosphopeptides, 38 phosphorylation sites in human NF-H were characterized. These include 33 KSP, lysine-threonine-proline (KTP) or arginine-serine-proline (RSP) sites and four unphosphorylated sites, all of which occur in the KSP repeat domain (residues 502-823); and one threonine phosphorylation site observed in a KVPTPEK motif. Six KSP sites were not characterized because of the failure to isolate and identify corresponding phosphopeptides. Heterogeneity in serine and threonine phosphorylation was observed at three sites or deduced to occur at three sites on the basis of enzyme specificity. As a result of the phosphorylated motifs identified (KSPAKEE, KSPVKEE, KS/TPEKAK, KSPEKEE, KSPVKAE, KSPAEAK, KSPPEAK, KSPEAKT, KSPAEVK, and KVPTPEK), human NF-H tail domain is postulated to be a substrate of proline-directed kinases. The threonine-phosphorylated KVPTPEK motif suggested the existence of a novel proline-directed kinase
— id: 24260, year: 1998, vol: 37, page: 3931, stat: Journal Article,

Mitogen-activated protein kinases (Erk1,2) phosphorylate Lys-Ser-Pro (KSP) repeats in neurofilament proteins NF-H and NF-M
Veeranna; Amin ND; Ahn NG; Jaffe H; Winters CA; Grant P; Pant HC
1998 Jun 1;18(11):4008-4021, Journal of neuroscience
Mammalian neurofilament proteins, particularly midsized (NF-M) and heavy (NF-H) molecular weight neurofilament proteins, are highly phosphorylated in axons. Neurofilament function depends on the state of phosphorylation of the numerous serine/threonine residues in these proteins. Most phosphorylation occurs in the lys-ser-pro (KSP) repeats in the C-terminal tail domains of NF-H and NF-M. In our previous study, cyclin-dependent kinase 5 (cdk5) was shown to phosphorylate specifically the KSPXK repeats in rat NF-H. Because 80% of the repeats are of the KSPXXXK type, it was of interest to determine which kinase phosphorylates these motifs. Using a synthetic KSPXXXK peptide to screen for a specific kinase, we fractionated rat brain extracts by column chromatography and identified extracellular signal-regulated kinase (Erk2) activated by an upstream activator, the mitogen-activated protein kinase kinase MAPKK (MEK), by Western blot analysis, sequence identification, and inhibition by a specific MEK inhibitor (PD 98059). The fraction containing Erk2, as well as bacterially expressed Erk1 and Erk2, phosphorylated all types of KSP motifs in peptides (KSPXK, KSPXXK, KSPXXXK, and KSPXXXXK) derived from NF-M and NF-H. They also phosphorylated an expressed 24 KSPXXXK repeat NF-H polypeptide, an expressed NF-H as well as dephosphorylated native rat NF-H, and NF-M proteins with accompanying decreases in their respective electrophoretic mobilities. A comparative kinetic study of Erk2 and cdk5 phosphorylation of KSPXK and KSPXXXK peptides revealed that, in contrast to cdk5, which phosphorylated only the KSPXK peptide, Erk2 could phosphorylate both. The preferred substrate for Erk2 was KSPXXXK peptide. The MEK inhibitor PD 98059 also inhibited phosphorylation of NF-H, NF-M, and microtubule-associated protein (MAP) in primary rat hippocampal cells and caused a decrease in neurite outgrowth, suggesting that Erk1,2 may play an important role in neurite growth and branching. These data suggest that neuronal Erk1 and Erk2 are capable of phosphorylating serine residues in diverse KSP repeat motifs in NF-M and NF-H
— id: 24259, year: 1998, vol: 18, page: 4008, stat: Journal Article,

Phosphorylation of human high molecular weight neurofilament protein (hNF-H) by neuronal cyclin-dependent kinase 5 (cdk5)
Pant AC; Veeranna; Pant HC; Amin N
1997 Aug 15;765(2):259-266, Brain research
Neurofilaments (NFs), the neuron-specific intermediate (i.e. approximately 10-nm diameter) filaments are major cytoskeletal components of most neurons. In a mature mammalian neuron, NFs are co-assembled from three subunits, NF-L (low), NF-M (medium), and NF-H (high), with molecular masses of 68, 95, and 115 kDa, respectively. Neurofilament proteins (NF-Ps), particularly, NF-H, are most extensively phosphorylated in large myelinated axons under normal conditions. This phosphorylation occurs on the serine residues of the lysine (Lys)-serine (Ser)-proline (Pro) (KSP) multiple amino acid repeats of the carboxy-terminal tail domain. Phosphorylation of KSP motifs affects physical, biochemical, and immunological properties of NF-H. For example, phosphorylation is thought to play a pivotal role in the maintenance of the neuronal cytoskeletal structure which influences the conduction velocity of the nerve fiber. The key components responsible for phosphorylation are not known. In this study, an identified cyclin-dependent kinase 5 (cdk5), isolated from nervous tissue, has been shown to phosphorylate the human NF-H (hNF-H) and affects its electrophoretic mobility. On the basis of the following observations, it is suggested that neuronal cdk5 (cdk5) phosphorylates KSPXK motifs in the human high molecular weight neurofilament (hNF-H) and affects its electrophoretic mobility. (1) A 14-mer synthetic peptide (KSPEKAKSPVKEEA) derived from hNF-H; (2) a bacterially expressed protein containing 14 KSPXK multiple repeats of hNF-H in C-terminal tail domain; and (3) a dephosphorylated hNF-H in neurofilament preparation are phosphorylated by cdk5. The decrease in molecular mass of hNF-H caused by dephosphorylated was completely recovered upon cdk5 phosphorylation. It is proposed that neuronal cdk5 regulates phosphorylation of the KSPXK motif in hNF-H and other cytoskeletal proteins with similar motifs in the nervous system
— id: 24261, year: 1997, vol: 765, page: 259, stat: Journal Article,

Expression of p67 (Munc-18), Cdk5, P-NFH and syntaxin during development of the rat cerebellum
Veeranna; Grant P; Pant HC
1997 ;19(2):172-183, Developmental neuroscience
Cyclin-dependent kinases (cdks), which regulate the cell division cycle, have also been found in postmitotic neurons. Cdk5, isolated from neural tissue, has been shown to phosphorylate neurofilaments (NFs). Instead of cyclins, however, other neuron-specific activators of cdk5 have been identified including a 67-kD protein (p67) which is identical to a syntaxin-binding protein (n-sec-1, Munc 18) that is thought to play a role in synaptic vesicle trafficking and transmitter release. These functions for p67 are not mutually exclusive since regulation of edk5 phosphorylation of cytoskeletal proteins may modulate axonal dynamics during growth, synaptogenesis and vesicle transport. To gain further insight into the role of p67 in neural tissue, we carried out a Western blot and immunohistochemical analysis of the developing rat cerebellum using antibodies to cdk5, p67, syntaxin and phosphorylated and nonphosphorylated neurofilaments. We assumed that spatiotemporal colocalizations of antigens might correlate with proposed functions for p67. The immunoblots showed that all antigens were developmentally regulated, and increased in expression from PN2 to the adult, with p67 and cdk5 showing a close temporal correlation. Immunohistochemically, p67 colocalized with cdk5 and P-NFH in selected fiber tracts, particularly those in the deep cerebellum. For the most part, p67 also showed strong colocalization patterns with syntaxin in regions of synaptogenesis throughout development such as the molecular layer and glomeruli of the inner nuclear layer. Finally, certain fiber tracts, the afferent fibers, climbing and mossy fibers and particularly the basket cell fibers that envelop and innervate Purkinje cell somata and dendrites, displayed colocalization of cdk5 and P-NFH without expressing any p67. Given the limitations of colocalization data in defining functional relationships, the results are consistent with the hypothesis that p67 is a multifunctional protein, its activity during cerebellum development dependent upon the neuronal phenotype, its location and its state of developmental maturation
— id: 24262, year: 1997, vol: 19, page: 172, stat: Journal Article,

Targeted disruption of the cyclin-dependent kinase 5 gene results in abnormal corticogenesis, neuronal pathology and perinatal death
Ohshima T; Ward JM; Huh CG; Longenecker G; Veeranna; Pant HC; Brady RO; Martin LJ; Kulkarni AB
1996 Oct 1;93(20):11173-11178, Proceedings of the National Academy of Sciences of the United States of America
Although cyclin-dependent kinase 5 (Cdk5) is closely related to other cyclin-dependent kinases, its kinase activity is detected only in the postmitotic neurons. Cdk5 expression and kinase activity are correlated with the extent of differentiation of neuronal cells in developing brain. Cdk5 purified from nervous tissue phosphorylates neuronal cytoskeletal proteins including neurofilament proteins and microtubule-associated protein tau in vitro. These findings indicate that Cdk5 may have unique functions in neuronal cells, especially in the regulation of phosphorylation of cytoskeletal molecules. We report here generation of Cdk5(-/-) mice through gene targeting and their phenotypic analysis. Cdk5(-/-) mice exhibit unique lesions in the central nervous system associated with perinatal mortality. The brains of Cdk5(-/-) mice lack cortical laminar structure and cerebellar foliation. In addition, the large neurons in the brain stem and in the spinal cord show chromatolytic changes with accumulation of neurofilament immunoreactivity. These findings indicate that Cdk5 is an important molecule for brain development and neuronal differentiation and also suggest that Cdk5 may play critical roles in neuronal cytoskeleton structure and organization
— id: 24263, year: 1996, vol: 93, page: 11173, stat: Journal Article,

Inhibition of neuronal cyclin-dependent kinase-5 by staurosporine and purine analogs is independent of activation by Munc-18
Veeranna; Shetty KT; Amin N; Grant P; Albers RW; Pant HC
1996 May;21(5):629-636, Neurochemical research
Neuronal cdk5 can phosphorylate certain lys-ser-pro (KSP) motifs of neurofilaments and tau protein in the nervous system. We have immunoprecipitated the cdk5 from rat brain using a polyclonal antibody raised against the C-terminus of cdk5. The immunoprecipitate has phosphorylated a KSPXK peptide analog of NF-H, as well as histone H1 and a bacterially expressed rat NF-H protein. The kinase activity was inhibited by staurosporine, isopentanyladenine and olomoucine in a dose dependent manner. Kinetic studies indicated Ki values of 39 nM, 38 microM and 8 microM, respectively for staurosporine, isopentanyladenine and olomoucine. The inhibition by staurosporine was non-competitive with respect to phosphoryl acceptor acceptor substrates. Western blot analysis of the immunoprecipitate showed both cdk5 and p67 (Munc-18), a putative regulator molecule of the kinase. Addition of p67 fusion protein enhanced the kinase activity of the immunoprecipitate by 60% above the basal activity. P67 elevated Ki values for both staurosporine and olomoucine. The degree of inhibition at high concentrations of these inhibitors was unaltered by exogenous p67 indicating a lack of competitive interactions with p67. The high affinity of staurosporine for cdk5 suggests that cdk5 may be one of the targets for the neurotropic effect of staurosporine
— id: 24264, year: 1996, vol: 21, page: 629, stat: Journal Article,

Neurofilament phosphorylation
Pant HC; Veeranna
1995 Sep-Oct;73(9-10):575-592, Biochemistry & cell biology
Neurofilament proteins (NFPs) are highly phosphorylated molecules in the axonal compartment of the adult nervous system. The phosphorylation of NFP is considered an important determinant of filament caliber, plasticity, and stability. This process reflects the function of NFs during the lifetime of a neuron from differentiation in the embryo through long-term activity in the adult until aging and environmental insult leads to pathology and ultimately death. NF function is modulated by phosphorylation-dephosphorylation in each of these diverse neuronal states. In this review, we have summarized some of these properties of NFP in adult nervous tissue, mostly from work in our own laboratory. Identification of sites phosphorylated in vivo in high molecular weight NFP (NF-H) and properties of NF-associated and neural-specific kinases phosphorylating specific sites in NFP are described. A model to explain the role of NF phosphorylation in determining filament caliber, plasticity, and stability is proposed
— id: 24265, year: 1995, vol: 73, page: 575, stat: Journal Article,

Neuronal cyclin-dependent kinase-5 phosphorylation sites in neurofilament protein (NF-H) are dephosphorylated by protein phosphatase 2A
Veeranna; Shetty KT; Link WT; Jaffe H; Wang J; Pant HC
1995 Jun;64(6):2681-2690, Journal of neurochemistry
Neurofilament (NF) protein [high molecular mass (NF-H)] is extensively phosphorylated in vivo. The phosphorylation occurs mainly in its characteristic KSP (Lys-Ser-Pro) repeat motifs. There are two major types of KSP motifs in the NF-H tail domain: KSPXKX and KSPXXX. Recent studies by two different laboratories have demonstrated the presence of a cdc2-like kinase [cyclin-dependent kinase-5 (cdk5)] in nervous tissue that selectively phosphorylates KSPXKX and XS/TXK motifs in NF-H and lysine-rich histone (H1). This article describes the identification of phosphatases dephosphorylating three different substrates: histone (H1), NF-H in a NF preparation, and a bacterially expressed C-terminal tail domain of NF-H, each containing KSPXKX repeats phosphorylated in vitro by cdk5. Among various phosphatases identified, protein phosphatase (PP) 2A from rabbit skeletal muscle appeared to be the most effective phosphatase in in vitro assays. Three phosphatase activity peaks--P1, P2, and P3--were partially purified from frozen rat spinal cord by ion exchange and size exclusion column chromatography and then characterized on the basis of biochemical, pharmacological, and immunochemical studies. One of the three peaks was identified as PP2A, whereas the others were mixtures of both PP2A and PP1. These three peaks could dephosphorylate cdk5-phosphorylated 32P-histone (H1), 32P-NF-H in the NF preparation, and 32P-NF-H tail fusion protein. These studies suggest the involvement of PP2A or a PP2A-like activity in the regulation of the phosphorylation state of KSPXKX motifs in NF-H
— id: 24266, year: 1995, vol: 64, page: 2681, stat: Journal Article,

Brain and spinal cord kappa opiate receptors and pharmacological responses to U-50,488H in rats of differing ages
Bhargava HN; Matwyshyn GA; Reddy PL; Veeranna
1994 May;48(1):87-91, Pharmacology biochemistry & behavior
The analgesic and hypothermic responses to U-50,488H (25 mg/kg IP), a kappa opiate receptor agonist, were determined in male Sprague-Dawley rats aged 4, 8, and 24 weeks. In addition, the characteristics of the binding of [3H]ethylketocylazocine (EKC) to kappa opiate receptors in whole brain and spinal cord of rats of three age groups were also determined. Administration of U-50,488H produced an age-related increase in the analgesic response in the rat, i.e., the older rats exhibited a higher intensity of analgesic response than the younger rats. U-50,488H also produced a hypothermic response. The response in 4- and 24-week-old rats was similar, but that in 8-week-old rats was smaller than the rats in the other two age groups. [3H]EKC bound to whole brain and spinal cord membranes of rats at a single high affinity site. The Bmax value of [3H]EKC in the brain and spinal cord of 24-week-old rats was significantly lower than in 4- and 8-week-old rats; however, the Kd values did not differ. It is concluded that kappa opiate receptor agonist produces age-related increase in its analgesic response and that such effects are not related to the characteristics of kappa receptors in the brain and spinal cord
— id: 24267, year: 1994, vol: 48, page: 87, stat: Journal Article,

Effect of morphine tolerance and abstinence on the binding of [3H]naltrexone to discrete brain regions and spinal cord of the rat
Reddy PL; Veeranna; Matwyshyn GA; Thorat SN; Bhargava HN
1994 Mar;25(2):355-361, General pharmacology
1. The effect of morphine tolerance and abstinence on the binding of [3H]naltrexone to discrete brain regions and spinal cord of the rat was determined. 2. Male Sprague-Dawley rats were implanted s.c. under light ether anesthesia with six morphine pellets for a 7-day period. Each pellet contained 75 mg of morphine base. Rats implanted with six placebo pellets each served as controls. 3. This procedure resulted in the development of tolerance to morphine as evidenced by decreased analgesic response to various doses of morphine. 4. The binding characteristics (Bmax or Kd values) of [3H]naltrexone, an opiate receptor antagonist, were determined in various tissues of morphine tolerant and abstinent rats. In the tolerant rats, the pellets were left in place at the time of sacrificing, whereas in the abstinent rats, the pellets were removed 18 hr prior to sacrificing. 5. The binding of [3H]naltrexone to opiate receptors on membranes prepared from brain regions (hypothalamus, hippocampus, cortex, pons and medulla, midbrain, corpus striatum and amygdala) and spinal cord of rats from various treatment groups was determined. 6. [3H]Naltrexone bound to tissue membranes at a single high affinity binding sites. The Bmax values of [3H]naltrexone to bind to opiate receptors on the membranes of amygdala and striatum were increased significantly in morphine tolerant rats when compared to the placebo controls, but the Kd values did not differ. 7. The Bmax and Kd values of [3H]naltrexone did not differ in any other brain region or spinal cord of morphine tolerant rats and their placebo controls. The binding constants of [3H]naltrexone were unaffected in morphine abstinent rats. 8. Previously we had shown that the binding of [3H]D-Ala2, MePhe4, Gly-ol5 enkephalin (DAMGO), a highly specific agonist for mu-opiate receptors was decreased in cortex, pons and medulla and spinal cord of morphine tolerant but not in the abstinent rats. In addition, delta and kappa receptors are unaffected in morphine tolerant and abstinent rats. 9. The results suggest that direction of change, as well as, the brain areas for mu-agonist and -antagonist opiate binding sites are affected differentially in morphine tolerant rats
— id: 24268, year: 1994, vol: 25, page: 355, stat: Journal Article,

Effects of naltrexone on the binding of [3H]D-Ala2, MePhe4, Gly-ol5-enkephalin to brain regions and spinal cord and pharmacological responses to morphine in the rat
Bhargava HN; Matwyshyn GA; Reddy PL; Veeranna
1993 Nov;24(6):1351-1357, General pharmacology
1. The effects of naltrexone pellet implantation and removal on the analgesic and hypothermic effects of morphine and the binding of 3H-D-Ala2, MePhe4, Gly-ol5-enkephalin (DAMGO) to mu-opiate receptors in rat brain regions and spinal cord were determined. 2. Male Sprague-Dawley rats were implanted subcutaneously with a pellet containing 10 mg of naltrexone for 7 days. Placebo pellet implanted rats served as controls. The pellets were removed on day 8, and the analgesic and hyperthermic effects were determined in the rat 24 hr later. Morphine produced a dose-dependent analgesic and hyperthermic responses in rats implanted with placebo pellets. Enhanced analgesic and hyperthermic responses to morphine were produced in rats implanted with naltrexone pellets. 3. The binding constants (Bmax and Kd values) of [3H]DAMGO in regions of the brain (amygdala, hypothalamus, striatum, midbrain, hippocampus, pons + medulla and cortex), and spinal cord of rats with naltrexone pellet left intact or removed were determined. The Bmax values of [3H]DAMGO were increased in all brain regions and spinal cord of rats in which the naltrexone pellets were left in place or removed prior to sacrificing. However, the Kd values of [3H]DAMGO were unaffected by naltrexone treatment. 4. It is concluded that enhanced analgesic and hyperthermic response to morphine is produced in rats implanted with naltrexone pellets and such alterations in the pharmacological responses are due to up-regulation of mu-opiate receptors in all the brain regions and spinal cord. Additionally whether the pellets were left intact (receptors blocked) or removed (receptors not blocked), the mu-opiate receptors were up-regulated in spinal cord and all the regions of the brain
— id: 24269, year: 1993, vol: 24, page: 1351, stat: Journal Article,

Evidence for the behavioral supersensitivity of dopamine D2 receptors without receptor up-regulation in morphine-abstinent rats
Reddy PL; Veeranna; Thorat SN; Bhargava HN
1993 Apr 2;607(1-2):293-300, Brain research
The effect of morphine tolerance-dependence and abstinence on the characteristics of dopamine D2 receptors in brain regions and spinal cord was determined in the rat. Male Sprague-Dawley rats were implanted s.c. under light ether anesthesia with 6 morphine pellets for a 7-day period, each containing 75 mg of morphine free base. Rats implanted with placebo pellets served as controls. This procedure resulted in the development of tolerance to morphine as evidenced by decreased analgesic response to a challenge dose of morphine. Similarly, the development of physical dependence was evidenced by a decrease in body weight and colonic temperature after morphine pellet removal (withdrawal). The binding characteristics (Bmax and Kd values) of [3H]spiroperidol to dopamine D2 receptors were determined in the tissues of morphine-tolerant and morphine-abstinent rats. In the tolerant rats, the pellets were left intact at the time of sacrificing, whereas, in the abstinent rats the pellets were removed 18 h prior to sacrificing. The binding of [3H]spiroperidol was determined in membranes prepared from brain regions (hypothalamus, hippocampus, cortex, pons and medulla, midbrain, corpus striatum and amygdala) and spinal cord of rats from various treatment groups. [3H]Spiroperidol bound to brain regions and spinal cord at a single high affinity site. The Bmax or the Kd values in brain regions and spinal cord of morphine-tolerant and -abstinent rats did not differ from their respective placebo controls. The behavioral responses to a selective dopamine D2 receptor agonist, 2-bromo-alpha-ergocryptine were also determine in the morphine-abstinent rats.(ABSTRACT TRUNCATED AT 250 WORDS)
— id: 24272, year: 1993, vol: 607, page: 293, stat: Journal Article,

Biochemical and behavioral studies on the interaction between mu- and kappa-opiate agonists in mice
Thorat SN; Veeranna; Reddy PL; Bhargava HN
1993 Jul 2;615(2):191-198, Brain research
Male Swiss-Webster mice were rendered tolerant to morphine by subcutaneous implantation of a morphine pellet, each containing 75 mg morphine base, for 3 days. Mice implanted with placebo pellets served as controls. A high degree of tolerance to the analgesic effect of morphine developed as evidenced by decreased analgesic response to various doses of morphine. A selective kappa-opiate agonist, U-50,488H (8, 16 and 32 mg/kg, i.p.) produced dose-dependent analgesic and hypothermic effects in mice implanted with placebo pellets. A significant decrease in the analgesic and hypothermic effects of U-50,488H was observed in morphine tolerant mice as compared to placebo-treated mice. Mice were rendered tolerant to U-50,488H by injecting the drug (25 mg/kg, i.p.) twice daily for 4 days. Vehicle injected mice served as controls. Tolerance to the analgesic and hypothermic effects of U-50,488H in mice injected chronically with the drug was evidenced by the decreases in the intensity of these responses when compared to those observed in vehicle injected controls. Morphine produced a dose-dependent analgesic and hypothermic effects in mice injected chronically with vehicle but the intensity of these effects was significantly lower in mice injected chronically with U-50,488H. These results indicate that a substantial tolerance to analgesic and hypothermic effects of U-50,488H develops in morphine tolerant mice. The effect of chronic injections of U-50,488H on the binding of [3H]ethylketocyclazocine (EKC) and [3H]D-Ala2,MePhe4,Gly-ol5-enkephalin (DAMGO) to whole brain and spinal cord kappa- and mu-opiate receptors was determined.(ABSTRACT TRUNCATED AT 250 WORDS)
— id: 24270, year: 1993, vol: 615, page: 191, stat: Journal Article,

Effect of chronic administration of U-50,488H, a kappa-opioid receptor agonist, on central dopamine D2 receptors of the rat
Veeranna; Reddy PL; Bhargava HN
1993 Apr 22;235(1):23-30, European journal of pharmacology
The effect of U-50,488H, a kappa-opiate agonist, induced tolerance and abstinence on the characteristics of dopamine D2 receptors of brain regions and spinal cord was determined in the rat. Male Sprague-Dawley rats were injected with U-50,488H (25 mg/kg i.p) or its vehicle twice a day for 4 days. This procedure resulted in the development of tolerance to the analgesic activity of U-50,488H. The binding characteristics (Bmax and Kd values) of [3H]spiroperidol to dopamine D2 receptors were determined in discrete brain regions (hypothalamus, hippocampus, cortex, pons and medulla, midbrain, corpus striatum and amygdala) and spinal cord of U-50,488H-tolerant and -abstinent rats. Rats labeled as tolerant to U-50,488H were injected with U-50,488H on day 5 and killed 1 h later, whereas those labeled abstinent were killed on day 5 without the injection of the drug. Vehicle-injected rats served as controls. [3H]Spiroperidol bound to brain regions and spinal cord membranes at a single high affinity site. The Bmax and Kd values of [3H]spiroperidol in brain regions and spinal cord of U-50,488H-tolerant and abstinent rats did not differ from their respective vehicle-injected controls. The behavioral responses (total distance travelled, horizontal activity, movement time, total number of movements, number of stereotypic movements, stereotypic time and rest time) to different doses of a selective dopamine D2 receptor agonist, 2-bromo-alpha-ergocryptine (bromocriptine) were also determined in rats treated chronically with U-50,488H.(ABSTRACT TRUNCATED AT 250 WORDS)
— id: 24271, year: 1993, vol: 235, page: 23, stat: Journal Article,

Phosphatase activity against neurofilament proteins from bovine spinal cord: effect of aluminium and neuropsychoactive drugs
Shetty KT; Veeranna; Guru SC
1992 Mar 16;137(1):83-86, Neuroscience letters
Protein phosphatase activity associated with neurofilament (NF) rich (Triton X-100 insoluble) fraction was extracted and partially characterised by using known inhibitors of protein phosphatases such as vanadate and fluoride. Protein phosphatase activity was demonstrated with reference to the dephosphorylation of endogenous substrate, NF protein and exogenous protein substrates, casein and phosvitin. Phosphoamino acids and beta-glycerophosphate were found to be poor substrates. Further, new observations have been made regarding the in vitro inhibitory effect of aluminium and the differential effects of some of the neuropsychoactive drugs. The findings could possibly lead to studies explaining the biochemical basis of aluminium induced neurotoxicity as well as the side effects associated with the long term medication of neuropsychoactive drugs
— id: 24273, year: 1992, vol: 137, page: 83, stat: Journal Article,

Phosphoserine phosphatase of human brain: partial purification, characterization, regional distribution, and effect of certain modulators including psychoactive drugs
Veeranna; Shetty KT
1990 Dec;15(12):1203-1210, Neurochemical research
Phosphoserine phosphatase (PSPase), a cytosolic enzyme has been purified 106 fold from human brain, by employing conventional protein purification techniques. The use of MgCl2 (10 mM) and chloroform treatment, during purification enabled the removal of non-specific proteins. The final enzyme preparation exhibited a broad pH optimum of 5.6-6.6 and could dephosphorylate both L and D enantiomers of the phosphoserine, but with different Km values for O-P-L serine (3.6 x 10(-5) M) and O-P-D serine (1 x 10(-4) M). Enzyme activity was found to be specific for phosphoserine, whereas other phosphoesters including phosphothreonine and phosphoproteins such as casein and phosvitin were found to be poor substrates. The enzyme activity was uncompetitively inhibited by L-serine. Further the PSPase activity was inhibited by vanadate, (41%), trifluoperazine (23%), chlorpromazine (34%) at an equimolar concentration of 1 mM, whereas lithium and ethanol did not influence the enzyme activity. Minor tranquilizers such as diazepam and chlordiazepoxide activated the enzyme activity to an extent of 13% and 59% respectively. In addition, species and regionwise heterogeneity was observed with respect to distribution of enzyme activity in six major areas of human, rabbit and rat brains
— id: 24274, year: 1990, vol: 15, page: 1203, stat: Journal Article,