We used Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) to detect the expression of the central and peripheral cannabinoid receptor (CB1 and CB2 respectively) mRNA and western blotting to show the presence of the CB1 protein in subregions of the human eye. We found that CB2 mRNA transcripts were undetectable, while the levels of CB1 mRNA were significantly expressed in the human retina (25.8% ± 2.46), ciliary body (210% ± 11.55) and iris (62.7% ± 5.94) when compared to those of the normalizing reference gene β2 microglobulin (β2m). The CB1 gene encodes a functional protein which is detected in its glycosylated (63 kDa) and unglycosylated (54 kDa) form in the same areas by a specific purified antibody raised against the amino terminus (residues 1-77) of the CB1 receptor. These results gave further strength to the proposed role of the CB1 receptor in controlling intraocular pressure (IOP) and sustained the proposed antiglaucoma properties of marijuana. To confirm that the CB1 peptide was functionall...

Progression from drugs such as alcohol and tobacco to marijuana and 'harder' drugs is common in adolescents. To assess whether progression occurs in an adolescent rodent model, we dosed rats with 5% or 10% alcohol (EtOH), or water under pair-watered or free intake conditions, from PD21 to PD70 [designated PPA, pre- and peri-adolescent]. Fluid was available 1 hr daily, and both 5% and 10% EtOH induced high SECs. After dosing was concluded, we assessed carryover effects to later cocaine administration for three potential modes of carryover. Direct carryover effects were seen: PPA 5% EtOH significantly increased adult IV cocaine self-administration, 5% and 10% decreased cocaine conditioned place preference (reported at Neuroscience '99), and 5% and 10% altered cocaine-stimulated motor activity in a gender-specific manner. Acute withdrawal carrover effects were not observed on tests of cocaine-induced activity and schedule-induced polydipsia, with repeated testing during withdrawal from alcohol. Drug-associate...

Spinocerebellar ataxia type 2 (SCA2) is an autosomal dominantly inherited, neurodegenerative disease caused by an expansion of polyglutamine tracts in the cytosolic protein ataxin-2 (Atx2). Cerebellar Purkinje cells (PCs) are predominantly affected in SCA2. The cause of PC degeneration in SCA2 is unknown. Here we demonstrate that mutant Atx2–58Q, but not wild-type (WT) Atx2–22Q, specifically associates with the cytosolic C-terminal region of type 1 inositol 1,4,5-trisphosphate receptor (InsP3R1), an intracellular calcium (Ca2+) release channel. Association with Atx2–58Q increased the sensitivity of InsP3R1 to activation by InsP3 in planar lipid bilayer reconstitution experiments. To validate physiological significance of these findings, we performed a series of experiments with an SCA2–58Q transgenic mouse model that expresses human full-length Atx2–58Q protein under the control of a PC-specific promoter. In Ca2+ imaging experiments, we demonstrated that stimulation with 3,5-dihydroxyphenylglycine (DHPG) r...

Endocannabinoids (eCBs) are retrograde neurotransmitters that modulate the function of many types of synapses. The presence of eCBs, their CB1 receptor (CB1R), and metabolizing enzymes at embryonic and early postnatal periods have been linked to developmental processes such as neuronal proliferation, differentiation, and migration, axon guidance, and synaptogenesis. Here, we demonstrate the presence of a functional eCB system in the developing visual system and the role of CB1R during axon growth and retinothalamic development. Pharmacological treatment of retinal explants and primary cortical neuron cultures with ACEA, a selective CB1R agonist, induced a collapse of the growth cone (GC). Furthermore the application of AM251, a CB1R inverse agonist, to the neuronal cultures increased the surface area of GC. In vivo , intraocular injection of ACEA diminished retinal projection growth, while AM251 promoted growth and caused aberrant projections. In addition, compared with their wild-type littermates, CB1R-de...

In mammals, potentially dangerous signals from the environment are detected by neurons in the pain pathway. Specialized neurons within the peripheral nervous system called nociceptive neurons sense noxious chemical, thermal, or mechanical stimuli and transmit signals to the CNS. It is well known

The endocannabinoid system is emerging as an integral component in central and peripheral regulation of feeding and energy balance. Our investigation analyzed behavioral roles for cannabinoid mechanisms of the pontine parabrachial nucleus (PBN) in modulating intake of presumably palatable foods containing fat and/or sugar. The PBN serves to gate neurotransmission associated with, but not limited to, the gustatory properties of food. Immunofluorescence and in vitro [35S]GTPγS autoradiography of rat tissue sections containing the PBN revealed the presence of cannabinoid receptors and their functional capability to couple to their G-proteins after incubation with the endocannabinoid 2-arachidonoyl glycerol (2-AG). The selective cannabinoid 1 receptor (CB1R) antagonist AM251 [ N -(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1 H -pyrazole-3-carboxamide] prevented the response, demonstrating CB1R mediation of 2-AG-induced coupling. Microinfusions of 2-AG into the PBN in behaving rats robustl...

Racism is a threat to public health. Race is a sociopolitical construct that has been used for generations to create disparities in educational access, housing conditions, exposure to environmental contaminants, and access to health care. Collectively, these disparities have a negative impact on the health of non-white Americans. The National Institutes of Health (NIH) funds biomedical research, including basic neuroscience research, aimed at understanding the mechanisms and consequences of health and disease in Americans. NIH has recently acknowledged its own structural racism, the disadvantage this perpetuates in the biomedical research enterprise, and has announced its commitment to eliminating these disparities. Here, we discuss different rates of disease in U.S. citizens from different racial backgrounds. We next describe ways in which the biomedical research enterprise (1) has contributed to health disparities and (2) can contribute to the solving this problem. Based on our own scientific expertise, ...

P/Q-type Ca2+ currents through presynaptic CaV2.1 channels initiate neurotransmitter release, and differential modulation of these channels by neuronal calcium-binding proteins (nCaBPs) may contribute to synaptic plasticity. The nCaBPs calcium-binding protein 1 (CaBP1) and visinin-like protein-2 (VILIP-2) differ from calmodulin (CaM) in that they have an N-terminal myristoyl moiety and one EF-hand that is inactive in binding Ca2+. To determine whether myristoylation contributes to their distinctive modulatory properties, we studied the regulation of CaV2.1 channels by the myristoyl-deficient mutants CaBP1/G2A and VILIP-2/G2A. CaBP1 positively shifts the voltage dependence of CaV2.1 activation, accelerates inactivation, and prevents paired-pulse facilitation in a Ca2+-independent manner. Block of myristoylation abolished these effects, leaving regulation that is similar to endogenous CaM. CaBP1/G2A binds to CaV2.1 with reduced stability, but in situ protein cross-linking and immunocytochemical studies revea...

Endocannabinoid (eCB)-mediated forms of long-term synaptic plasticity occur in several brain regions, but much remains unknown about their basic properties and underlying mechanisms. Here, we present evidence that eCB-mediated long-term depression (eCB-LTD) at excitatory synapses on medium spiny neurons in the striatum requires presynaptic activity coincident with CB1 receptor activation. This dual requirement for CB1 activation and presynaptic activity is a mechanism by which eCB-LTD may be made synapse specific.