Quantitative histochemical study on the postnatal development of perineuronal nets in the retrosplenial cortex of albino rats.

摘要

The study was conducted to demonstrate the patterns of development of perineuronal nets (PNs), and to determine the histochemical and morphometric data on PNs in the retrosplenial cortex (RSC). 122 Wistar rats at birth (n=6), and postnatal weeks 1 (PW1; n=10), 2 (n=10), 3 (n=10), 4 (n=12), 5 (n=12), 6 (n=12), 7 (n=12), 8 (n=12), 10 (n=12), as well as 12 weeks old animals (n=12) were perfused transcardially under deep ether anaesthesia with saline, followed with 4% paraformaldehyde in 0.1M sodium cacodylate buffer. The brains were isolated and 2-3 mm thick tissue slices traversing the granular and agranular subfields of retrosplenial cortex were cut and fixed. The histochemical and quantitative characteristics of postnatal extracellular matrix (ECM) in RSC were observed with a cationic sulfated proteoglycans (PG) and with certain plant lectins from Vicia villosa agglutinin (VVA), Wisteria floribunda agglutinin (WFA), or Glycine max agglutinin (SBA) for the determination of the N-acetylgalactosamine containing glycoproteins (GPs). Results revealed that the differentiating PNs were observed as early as 1-2 weeks postnatally. At Pw 6-8, the overall density and intensity of labelled PNs progressively increased and reached the adult stage of development. The PNs also displayed differential labelling characteristics for the lectin or CIC double staining which were differentially identified into three types: PGs, GPs and PGs or GPs coats. Morphometric measurements revealed that the total density of net-associated retrosplenial neurons (RSNs) at Pw2 was ~1.80+or-0.29/UA after labelling with WFA/CIC double staining technique, and this variant increased polynomially with progression of postnatal age reaching 9.12+or-1.82/UA at Pw12. Quantitative analysis showed that the gain of net-associated RSNs increased rapidly during the first postnatal month, and then increased 1.73 fold during the second month and 1.02 at the third month. The numerical density of net associated RSNs significantly increased with increase in age. The numerical densities of PGs and GPs coated neurons, as well as those exhibiting coat complexes, increased polynomially with progression of animal age. The absolute numerical density of RSNs labelled with VVA, WFA, SBA, CIC, and CIC-Bodian staining measured ~2.12+or-0.17, 1.80+or-0.13, 0.00, 0.00 and 3.00+or-0.18/UA, respectively, at Pw2 and increased to 6.18+or-1.23, 5.00+or-1.03, 2.90+or-0.69, 3.85+or-0.76 and 4.30+or-1.13/UA at Pw12, respectively. The labelling indices of RSNs stained with VVA, WFA, SBA, CIC, and CIC-Bodian were 3.60, 2.94, 0.00, 0.00 and 3.52 at Pw2, respectively. It is indicated that the perineuronal ECM components are significantly correlated with age and suggest a possible developmental and biological significance including promotion of migration and functional maturation of net-associated neurons in the RSC.