A comprehensive analysis of the effect of DSP4 on the locus coeruleus noradrenergic system in the rat

Abstract

Degeneration of the noradrenergic neurons in the locus coeruleus (LC) is a major component of Alzheimer's (AD) and Parkinson's disease (PD), but the consequence of noradrenergic neuronal loss has different effects on the surviving neurons in the two disorders. Therefore, understanding the consequence of noradrenergic neuronal loss is important in determining the role of this neurotransmitter in these neurodegenerative disorders. The goal of the study was to determine if the neurotoxin N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP4) could be used as a model for either (or both) AD or PD. Rats were administered DSP4 and sacrificed 3 days 2 weeks and 3 months later. DSP4-treatment resulted in a rapid, though transient reduction in norepinephrine (NE) and NE transporter (NET) in many brain regions receiving variable innervation from the LC. Alpha(1)-adrenoreceptors binding site concentrations were unchanged in all brain regions at all three time points. However, an increase in alpha(2)-AR was observed in many different brain regions 2 weeks and 3 months after DSP4. These changes observed in forebrain regions occurred without a loss in LC noradrenergic neurons. Expression of synthesizing enzymes or NET did not change in amount of expression/neuron despite the reduction in NE tissue content and NET binding site concentrations at early time points, suggesting no compensatory response. In addition, DSP4 did not affect basal activity of LC at any time point in anesthetized animals, but 2 weeks after DSP4 there is a significant increase in irregular firing of noradrenergic neurons. These data indicate that DSP4 is not a selective LC noradrenergic neurotoxin, but does affect noradrenergic neuron terminals locally, as evident by the changes in transmitter and markers at terminal regions. However, since DSP4 did not result in a loss of noradrenergic neurons, it is not considered an adequate model for noradrenergic neuronal loss observed in AD and PD.

Figure 1

Weight of animals 3 days, 2 weeks and 3 months after DSP4 (50 mg/kg, ip) or saline. Animals that received DSP4 initially (3 days later) gained weight at a slower rate than control animals but quickly caught up to control at 2 weeks. * Indicates significant difference from saline-treated animals.

Figure 2

Norepinephrine (NE) and dopamine (DA) tissue content in prefrontal cortex (PFC), hippocampus (HP), locus coeruleus (LC), cerebellum (CB), amygdala (Amy) and septum/bed nucleus of the stria terminalis (sep/BNST) 3 days, 2 weeks and 3 months after administration of DSP4 and saline. Values are converted to percent control for each time point. NE is significantly reduced 3 days after DSP4 in the PFC, HP and sep/BNST and 2 weeks after DSP4 in HP and CB. NE tissue content is significantly elevated in the sep/BNST 2 weeks after DSP4. NE levels are normal 3 months after DSP4 in all regions. DA levels are only reduced in the HP 2 weeks after DSP4. * Indicates significant difference from saline-treated animals.

Figure 3

Norepinephrine transporter (NET) (³H-nisoxetine) binding site concentrations in forebrain regions 3 days, 2 weeks and 3 months after administration of saline or DSP4. NET binding site concentrations were measured in the frontal cortex (FC), septum (sep), bed nucleus of the stria terminalis (BNST), anterior ventricular thalamus (AVT), paraventricular hypothalamic nucleus (PVN), hippocampus (HP), habenula (Hab), hypothalamus (hypo), amygdala (Amy) and substantia nigra/ventral tegmental area (SN/VTA). Values are converted to percent saline for each time point. Three days after DSP4 NET binding site concentrations are significantly reduced in the FC and HP. Two weeks after DSP4 NET binding site concentrations are significantly reduced in the sep, AVT and Hab with normal levels in FC and HP. Three months after DSP4 NET binding site concentrations are similar to saline in all regions studied. * Indicates significant difference from saline treated animals.

Figure 4

Autoradiographic norepinephrine transporter (NET) binding in specific forebrain regions of saline and DSP4-treated animals 2 weeks after DSP4 administration. Arrows indicate NET specific binding in the frontal cortex (FC), septum (sep), anterior ventral thalamic nucleus (AVT), paraventricular nucleus (PVN), hippocampus (HP) and habenula (Hab).

Figure 5

α₂-Adrenoreceptor (α₂-AR) (³H-RX821002) binding sites concentrations in forebrain regions 3 days, 2 weeks and 3 months after administration of saline or DSP4. α₂-AR binding site concentrations were measured in the frontal cortex (FC), striatum, (str), anterior septum (Ant sep), ventral septum (Vent sep), bed nucleus of the stria terminalis (BNST), dorsal thalamic nucleus (DTN), paraventricular thalamic nucleus (PVTN), hippocampus (HP), hypothalamus (hypo), amygdala (Amy), substantia nigra/ventral tegmental area (SN/VTA), lateral posterior thalamic nucleus (LPT), central grey (CG). Values are converted to percent control for each time point. α₂-AR binding site concentrations in DSP4-treated animals are not different from saline-treated animals 3 days later, but 2 weeks after DSP4 α₂-AR binding site concentrations are significantly elevated in the DTN, PVNT and hypo. Three months after DSP4 α₂-AR binding site concentrations are significantly elevated in the Ant sep and Amy. * Indicates significant difference from saline-treated animals.

Figure 6

Autoradiographic α₂-adrenergic receptor (α₂-AR) binding in specific forebrain regions of saline and DSP4-treated animals 2 weeks after DSP4 administration. Arrows indicate specific α₂-AR binding in the frontal cortex (FC), septum (sep), paraventricular thalamic nucleus (PVNT), dorsal thalamic nucleus (DTN), amygdala (Amy) and hypothalamus (Hypo).