Prepuberal stimulation of 5-HT7-R by LP-211 in a rat model of hyper-activity and attention-deficit: permanent effects on attention, brain amino acids and synaptic markers in the fronto-striatal interface

The cross-talk at the prefronto-striatal interface involves excitatory amino acids, different receptors, transducers and modulators. We investigated long-term effects of a prepuberal, subchronic 5-HT7-R agonist (LP-211) on adult behaviour, amino acids and synaptic markers in a model for Attention-Deficit/Hyperactivity Disorder (ADHD). Naples High Excitability rats (NHE) and their Random Bred controls (NRB) were daily treated with LP-211 in the 5th and 6th postnatal week. One month after treatment, these rats were tested for indices of activity, non selective (NSA), selective spatial attention (SSA) and emotionality. The quantity of L-Glutamate (L-Glu), L-Aspartate (L-Asp) and L-Leucine (L-Leu), dopamine transporter (DAT), NMDAR1 subunit and CAMKIIα, were assessed in prefrontal cortex (PFC), dorsal (DS) and ventral striatum (VS), for their role in synaptic transmission, neural plasticity and information processing. Prepuberal LP-211 (at lower dose) reduced horizontal activity and (at higher dose) increased SSA, only for NHE but not in NRB rats. Prepuberal LP-211 increased, in NHE rats, L-Glu in the PFC and L-Asp in the VS (at 0.250 mg/kg dose), whereas (at 0.125 mg/kg dose) it decreased L-Glu and L-Asp in the DS. The L-Glu was decreased, at 0.125 mg/kg, only in the VS of NRB rats. The DAT levels were decreased with the 0.125 mg/kg dose (in the PFC), and increased with the 0.250 mg/kg dose (in the VS), significantly for NHE rats. The basal NMDAR1 level was higher in the PFC of NHE than NRB rats; LP-211 treatment (at 0.125 mg/kg dose) decreased NMDAR1 in the VS of NRB rats. This study represents a starting point about the impact of developmental 5-HT7-R activation on neuro-physiology of attentive processes, executive functions and their neural substrates

Prepuberal intranasal dopamine treatment in an animal model of ADHD ameliorates deficient spatial attention, working memory, amino acid transmitters and synaptic markers in prefrontal cortex, ventral and dorsal striatum

Intranasal application of dopamine (IN-DA) has been shown to increase motor activity and to release DA in the ventral (VS) and dorsal striatum (DS) of rats. The aim of the present study was to assess the effects of IN-DA treatment on parameters of DA and excitatory amino acid (EAA) function in prepuberal rats of the Naples high-excitability (NHE) line, an animal model for attention-deficit hyperactivity disorder (ADHD) and normal random bred (NRB) controls. NHE and NRB rats were daily administered IN-DA (0.075, 0.15, 0.30 mg/kg) or vehicle for 15 days from postnatal days 28–42 and subsequently tested in the Làt maze and in the Eight-arm radial Olton maze. Soluble and membrane-trapped l-glutamate (l-Glu) and l-aspartate (l-Asp) levels as well as NMDAR1 subunit protein levels were determined after sacrifice in IN-DA- and vehicle-treated NHE and NRB rats in prefrontal cortex (PFc), DS and VS. Moreover, DA transporter (DAT) protein and tyrosine hydroxylase (TH) levels were assessed in PFc, DS, VS and mesencephalon (MES) and in ventral tegmental area (VTA) and substantia nigra, respectively. In NHE rats, IN-DA (0.30 mg/kg) decreased horizontal activity and increased nonselective attention relative to vehicle, whereas the lower dose (0.15 mg/kg) increased selective spatial attention. In NHE rats, basal levels of soluble EAAs were reduced in PFc and DS relative to NRB controls, while membrane-trapped EAAs were elevated in VS. Moreover, basal NMDAR1 subunit protein levels were increased in PFc, DS and VS relative to NRB controls. In addition, DAT protein levels were elevated in PFc and VS relative to NRB controls. IN-DA led to a number of changes of EAA, NMDAR1 subunit protein, TH and DAT protein levels in PFc, DS, VS, MES and VTA, in both NHE and NRB rats with significant differences between lines. Our findings indicate that the NHE rat model of ADHD may be characterized by (1) prefrontal and striatal DAT hyperfunction, indicative of DA hyperactivty, and (2) prefrontal and striatal NMDA receptor hyperfunction indicative of net EAA hyperactivty. IN-DA had ameliorative effects on activity level, attention, and working memory, which are likely to be associated with DA action at inhibitory D2 autoreceptors, leading to a reduction in striatal DA hyperactivity and, possibly, DA action on striatal EAA levels, resulting in a decrease of striatal EAA hyperfunction (with persistence of prefrontal EAA hyperfunction). Previous studies on IN-DA treatment in rodents have indicated antidepressant, anxiolytic and anti-parkinsonian effects in relation to enhanced central DAergic activity. Our present results strengthen the prospects of potential therapeutic applications of intranasal DA by indicating an enhancement of selective attention and working memory in a deficit model

LC/MS/MS parameters.

<p>LC/MS/MS parameters.</p

Level of CaMKIIα protein measured by western blot in NHE (panel A) and NRB (panel B) rats after adolescent treatment with 0.125 or 0.250 mg/kg LP-211.

<p>Data are presented as % of vehicle response ± standard error.</p

Long term behavioural effects of prepuberal subchronic LP-211 treatment on activity and non selective attention in the Olton maze for adult NHE and NRB rats.

<p>The horizontal activity (A), the frequency of rearings (B) and their duration (C) after vehicle or treatment with 0.125, 0.250 or 0.500 mg/kg LP-211 are reported as mean ± SEM, (N = 6/group). For statistical analysis see the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0083003#s2" target="_blank">Results</a> section (* p<0.01).</p

Level of CaMKIIα protein measured in the PFC (panel A), DS (panel B) and VS (panel C) of NHE and NRB rats under basal conditions.

<p>Data are presented as mean ± standard error. For statistical analysis see the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0083003#s2" target="_blank">Results</a> section (* p<0.05).</p

Level of Glu/Leu (black) and Asp/Leu (white) expressed as ratio in PFC, DS and VS of NHE (panel A) and NRB (panel B) rats.

<p>Data after vehicle or treatment with 0.125 or 0.250/kg LP-211 are presented as mean ± SEM, (N = 6/group). For statistical analysis see the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0083003#s2" target="_blank">Results</a> section (* p<0.02).</p

Level of NMDAR1 protein measured by western blot in the forebrain sites of NHE (panel A) and NRB (panel B) rats after adolescent treatment with 0.125 or 0.250 mg/kg LP-211.

<p>Data are presented as % of vehicle response ± standard error. For statistical analysis see the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0083003#s2" target="_blank">Results</a> section (* p<0.05).</p

Long term behavioural effects of prepuberal subchronic LP-211 treatment on activity and non selective attention in the Làt maze for adult NHE and NRB rats.

<p>The frequency of corner crossings (A), the frequency of rearings (B), and their duration (C) after vehicle or treatment with 0.125, 0.250 or 0.500 mg/kg LP-211 are reported as mean ± SEM, (N = 6/group). For statistical analysis see the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0083003#s2" target="_blank">Results</a> section.</p

Level of DAT protein measured by western blot in the PFC, DS, VS and ME of NHE (panel A) and NRB (panel B) rats after adolescent treatment with 0.125 or 0.250 mg/kg LP-211.

<p>Data are given as % of vehicle response ± standard error. For statistical analysis see the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0083003#s2" target="_blank">Results</a> section (* p<0.05).</p