Coordinate regulation of tubulin and microtubule associated protein genes during development of hamster brain.

The present study documents the patterns of mRNA expression for 5 different tubulin genes and 4 of the structural microtubule-associated protein (MAP) genes during normal development of hamster forebrain. Northern blotting in conjunction with densitometric analysis was used to study changes in the levels of the mRNAs for alpha 1-tubulin, classes beta I-, beta II-, beta III- and beta IV-tubulin, as well as the mRNAs for tau, MAP1A, MAP1B and MAP2, using total RNA isolated from hamster forebrain at various embryonic (E) and postnatal (P) stages. Densitometric analyses of the autoradiograms from the Northern blots revealed that each of the tubulin genes exhibited distinct developmental patterns of expression, several of which appeared to be temporally correlated with the expression of specific MAP mRNAs. The beta I-, beta II- and beta III-tubulin mRNAs increased rapidly between late embryonic stages to birth, reached peak levels early in the first postnatal week, and declined thereafter. alpha 1-Tubulin mRNA was easily detected during embryonic stages, rose to peak levels at P7-P9 and then gradually declined. A similar pattern was seen for tau mRNA. After the first postnatal week, the size of the tau mRNA also shifted to a slightly larger size, presumably due to differential splicing. beta IV-Tubulin mRNA levels did not become significant until very late in postnatal development (3-4 weeks). MAP2 mRNA expression was unusual in that peak levels were reached at two different stages of development--an initial peak occurred in the first postnatal week, followed by a decline, and then a second rise occurred during the third and fourth postnatal weeks. The expression of the beta IV-tubulin mRNA coincided temporally with the second peak in MAP2 mRNA expression. MAP1B mRNA abruptly reached high levels at birth, remained abundant during the first two postnatal weeks, and then decreased. In contrast, MAP1A mRNA levels were low in the initial postnatal interval and increased only at very late developmental stages. The findings of a temporal correspondence in expression of high levels of tau and MAP1B with beta I-, beta II-, beta III- and alpha 1-tubulin mRNAs suggest that this profile of gene expression is one that endows greater plasticity to the neuronal cytoskeleton. Conversely, the transition to increased expression of beta IV-tubulin, MAP1A, and a larger tau mRNA species defines a portion of the molecular pattern that underpins the increased stability of neuronal form during maturation.

Immature corticospinal neurons respond to axotomy with changes in tubulin gene expression.

We have examined the expression of two different tubulin mRNAs in hamster corticospinal neurons that were axotomized at three different developmental stages; postnatal day 8 (P8), P20, and adult. In situ hybridization of histological sections of the sensorimotor cortex was done with 35S-labeled cDNA probes specific to alpha 1-tubulin and beta III-tubulin mRNAs at 2-14 days following unilateral transection of the corticospinal tract in the caudal medulla. Both film and emulsion autoradiography were used to detect changes in tubulin mRNA levels. Qualitative assessment indicated substantial decreases in both alpha 1-tubulin and beta III-tubulin mRNA levels in layer V neurons of the sensorimotor cortex following axotomy. The changes were apparent as early as 2 days postinjury for P20 and adult operates, but not for P8 operates. However, by 14 days postinjury, decreases in alpha 1-tubulin and beta III-tubulin gene expression were apparent in animals operated at all three developmental stages. These findings indicate that both immature and adult corticospinal neurons respond to axonal injury in a manner that is distinctly different from the peripheral neuron response.

Developmental patterns of intermediate filament gene expression in the normal hamster brain.

We have examined the patterns of expression of the major intermediate filament (IF) protein mRNAs during development of the hamster brain. Quantitative northern blotting was used to examine changes in the levels of mRNAs for the low, middle and high molecular weight neurofilament proteins (NF-L, NF-M, NF-H) as well as peripherin, vimentin and glial fibrillary acidic protein (GFAP). Total RNA was isolated from hamster brains at embryonic (E) days 12 and 14 and postnatal (P) days 1, 3, 5, 7, 9, 11, 13, 15, 20, 28 and 60-90 (adult), and probed with specific IF cDNAs. Northern blotting revealed that NF-L and NF-M mRNAs were present at very low levels in embryonic brain and that significant expression of these genes only occurred postnatally when the levels increased dramatically until P28 and then declined again in the adult. Increases in NF-H mRNA levels were somewhat delayed relative to those of NF-L and NF-M. NF-H mRNA was not seen at embryonic stages and was expressed at very low levels prior to P9; after that time the levels increased rapidly until P28 and then declined in the adult. Two of the type III IF genes, peripherin and vimentin, followed a pattern of expression opposite that of the NF genes. Both peripherin and vimentin mRNAs were present in embryonic brain and were expressed at higher levels during early postnatal stages than at later times. The magnitude and rate of reduction in vimentin gene expression in the postnatal interval was much greater than that of peripherin. GFAP mRNA levels were extremely low prior to P9 after which a robust increase occurred, followed by a decline in the adult. We discuss the implication of the dramatic changes in IF isotype expression in brain to the pathways of both neuronal and glial development in vivo.