Spatial memory is related to hippocampal subcellular concentrations of calcium-dependent protein kinase C isoforms in young and aged rats

Abstract

Relationships were examined between spatial learning and hippocampal concentrations of the alpha, beta2, and gamma isoforms of protein kinase C (PKC), an enzyme implicated in neuronal plasticity and memory formation. Concentrations of PKC were determined for individual 6-month-old (n = 13) and 24-month-old (n = 27) male Long-Evans rats trained in the water maze on a standard place-learning task and a transfer task designed for rapid acquisition. The results showed significant relationships between spatial learning and the amount of PKC among individual subjects, and those relationships differed according to age, isoform, and subcellular fraction. Among 6-month-old rats, those with the best spatial memory were those with the highest concentrations of PKCgamma in the particulate fraction and of PKCbeta2 in the soluble fraction. Aged rats had increased hippocampal PKCgamma concentrations in both subcellular fractions in comparison with young rats, and memory impairment was correlated with higher PKCgamma concentrations in the soluble fraction. No age difference or correlations with behavior were found for concentrations of PKCgamma in a comparison structure, the neostriatum, or for PKCalpha in the hippocampus. Relationships between spatial learning and hippocampal concentrations of calcium-dependent PKC are isoform-specific. Moreover, age-related spatial memory impairment is associated with altered subcellular concentrations of PKCgamma and may be indicative of deficient signal transduction and neuronal plasticity in the hippocampal formation.

Figure 1

(A) Representative image of standards prepared from pooled tissue of naive, young rats, ranging from 4–16 μg of total protein and immunostained for PKCγ. Each gel was loaded with standards in this range for either soluble or particulate fractions from either the hippocampus or the neostriatum. (B) Plot and linear regression of tissue standards in A. The abscissa represents the concentration of total protein loaded in each lane, and the ordinate represents the integrated measure of band optical density multiplied by the area in number of pixels. (C) Representative image of sample lanes immunostained for PKCγ. Each lane was loaded with 10 μg of total protein from an individual rat from either the soluble or the particulate fraction of either the hippocampus or the neostriatum. Samples were pseudorandomly loaded on the gel such that each group (young, aged-unimpaired, and aged-impaired) was represented once in each group of three lanes. Ten samples were run simultaneously with six standards on each gel, and sample values were extrapolated from the corresponding standard curve.

Figure 2

(A)Twenty-four-month-old rats (□) were significantly impaired in comparison with 6-month-old rats (○) during place learning. Aged rats (A, Inset) showed greater heterogeneity than young rats (Y). (B) There was no reliable difference between 24-month-old rats (□) and 6-month-old rats (○) during acquisition of the transfer task. During the probe trial (Inset), the subgroup of aged rats that performed outside the range of young rats on the original spatial task (AI, aged impaired; n = 15) also showed significantly less spatial bias than either aged rats that performed within the range of the young rats (AU, aged-unimpaired; n = 12) or young rats (Y, n = 13).

Figure 3

(A) There was a significant correlation between the learning index and the concentrations of PKCγ in the particulate fraction of young rats (r = −0.65, P < 0.05). Young rats with the lowest learning index scores, indicating the best spatial memory, had the highest concentrations of PKCγ in the particulate, or membrane-bound, fraction. (B) The learning index was correlated significantly with PKCβ₂ concentrations in the soluble fraction among young rats (r = −0.58, P < 0.05). Young rats with the lowest learning index scores had the highest concentrations of PKCβ₂ in the soluble fraction. (C) A significant correlation was found between the learning index and the concentrations of PKCγ in the soluble fraction of aged rats (r = 0.40, P < 0.05). Aged rats with the highest learning index scores, indicating the worst spatial memory, had the highest concentrations of PKCγ in the soluble, or cytosolic, fraction.