5-Hydroxytryptophan during critical postnatal period improves cognitive performances and promotes dendritic spine maturation in genetic mouse model of phenylketonuria

Abstract

Although phenylketonuria (PKU) is the most common genetic cause of mental retardation, the cellular mechanisms underlying impaired brain function are still unclear. Using PAHenu2 mice (ENU2), the genetic mouse model of PKU, we previously demonstrated that high phenylalanine levels interfere with brain tryptophan hydroxylase activity by reducing the availability of serotonin (5-hydroxytryptamine, 5-HT), crucial for maturation of neuronal connectivity in the prefrontal cortex (PFC), around the third postnatal week, a critical period for cortical maturation. 5-Hydroxytryptophan (5-HTP), the product of tryptophan hydroxylation, is known to be a better treatment to increase brain 5-HT levels. In this study we investigated the role of 5-HT during the early postnatal period in cognitive disturbances and in cortical dendritic alterations of PKU subjects by restoring temporarily (postnatal days 14-21) physiological brain levels of 5-HT in ENU2 through 5-HTP treatment. In adult ENU2 mice early 5-HTP treatment reverses cognitive deficits in spatial and object recognition tests accompanied by an increase in spine maturation of pyramidal neurons in layer V of the prelimbic/infralimbic area of the PFC, although locomotor deficits are not recovered by treatment. Taken together, our results support the hypothesis that mental retardation in PKU depends on reduced availability of brain 5-HT during critical developmental periods that interferes with cortical maturation and point to 5-HTP supplementation as a highly promising additional tool to heal PKU patients.

Fig. 1

5-HTP administration between postnatal days 14 and 21 improves cognitive performances in adult ENU2 mice. (a) Locomotor measures are shown as crossings (left panel) and distance moved (right panel). Insets show total responses over 1-h test. (b) Results obtained in the object recognition test (left panel) expressed as time spent exploring the two identical objects (A1, A2) during S2 pre-test session (middle panel) and as time spent exploring either the novel (B) or the familiar (A3) object during S3 test session (right panel). 5-HTP early treatment restored object recognition ability in ENU2 mice. (c) Results obtained in the spatial novelty test (right panel) expressed as mean time spent exploring all objects (A, B, C, D) during S2, S3 and S4 (middle panel), and as changes in the time spent exploring the displaced object (DO; A, B) and the non-displaced object (NDO; C, D) between the last habituation session (S4) and the test session (S5) (right panel). 5-HTP early treatment also restored spatial novelty discrimination in ENU2 mice. All data are expressed as mean±s.e.m. * p<0.05 vs. all other groups; ^#p<0.05 vs. familiar object; §p<0.05 vs. NDO.

Fig. 2

5-HTP administration between postnatal days 14 and 21 increases the number of mature dendritic spines in frontal cortical pyramidal neurons of adult ENU2 mice. (a) Schematic representation of prefrontal cortex (PFC), photomicrograph of representative Golgi–Cox impregnated medium pyramidal neuron of layer V of the PFC and schematic depiction of the neuron showing the position of apical shaft and basal and apical dendrites (left panel). There were no significant differences between groups in analysis of dendritic morphology: apical and basal dendritic length and basal branching (right panels). (b) High-power photomicrographs of the representative apical dendritic segment and of categories of spines considered (mature: M=mushroom; S=stubby; and immature: T=thin). Scale bar, 10 μm. ENU2 mice displayed significantly more reduced basal and apical dendrite density than all other groups. Although ENU2-Sal and ENU2-5-HTP mice displayed more basal and apical spine density than ENU2 mice, only ENU2-5-HTP mice showed a recovery of spine maturation to WT levels. All data are expressed as mean±s.e.m. * p<0.05, ** p<0.001.